Anti-atrophic role of Ononis natrix subsp. hispanica extract in skeletal muscle atrophy: Pathway-targeted insights and experimental validation

Abstract

Skeletal muscle atrophy, a condition associated with sarcopenia, cachexia, and various metabolic diseases, significantly impairs physical function and quality of life. This study investigates the potential of Ononis natrix subsp. hispanica water extract, rich in quercetin, luteolin, and apigenin, to mitigate muscle atrophy in an in vitro model. Atrophy was induced in differentiated C2C12 myotubes using dexamethasone, and the effects of the extract and a mixture of its phytochemicals (QLA mixture) were evaluated on myotube morphology and key protein degradation markers. The extract significantly reduced dexamethasone-induced myotube diameter loss from 28% to 13%–15.4% (p < 0.001). Similarly, the QLA mixture limited diameter loss to 14.6%, maintaining a morphology closer to the control. The extract and QLA mixture attenuated the dexamethasone-induced upregulation of muscle-specific ubiquitin ligases MAFbx and MuRF1, with mRNA levels reduced by 2.1–2.4-fold and 1.9–2.4-fold, respectively (p < 0.001). Western blot analysis validated the reduction in MAFbx protein levels. Molecular docking studies demonstrated robust binding interactions of quercetin, luteolin, and apigenin with TGFBR1 and IKBKB, showing binding energies as low as −10.1 kcal/mol. Notably, luteolin formed six hydrogen bonds with TGFBR1, while apigenin displayed four hydrogen bonds with IKBKB, underscoring their strong inhibitor potential. KEGG pathway enrichment analysis highlighted the involvement of PI3K-Akt, NF-κB, and FoxO signaling pathways, elucidating the extract's multi-target mechanisms. These findings establish the therapeutic promise of O. natrix extract as a natural agent for managing skeletal muscle atrophy and improving muscle health.