Targeting SOD3-Mediated Oxidative Stress with Betulonic Acid Protects Against Steroid-Induced Femoral Head Necrosis

Steroid-induced osteonecrosis of the femoral head (SONFH) is a devastating complication of glucocorticoid therapy, fundamentally driven by oxidative stress. Although Betulonic acid (Ba), a natural pentacyclic triterpenoid, exhibits potent antioxidant properties, its therapeutic efficacy in SONFH remains unknown. This study investigates the protective effects of Ba against SONFH, specifically focusing on its capacity to restore extracellular redox homeostasis via superoxide dismutase 3 (SOD3).

A rat SONFH model was induced using lipopolysaccharide and methylprednisolone, followed by Ba treatment. Femoral head architecture was evaluated via Micro-CT and histopathology. In vitro, MC3T3-E1 osteoblasts were treated with dexamethasone (Dex) and Ba. Cell viability, intracellular ROS, and superoxide anion levels were assessed. RNA sequencing identified differentially expressed genes, and the mechanism was validated using Sod3 siRNA knockdown and lentiviral overexpression assays.

Ba administration significantly attenuated osteonecrosis in vivo, preserving bone mineral density (p < 0.01) and preventing trabecular collapse. In vitro, Ba mitigated Dex-induced osteoblast cytotoxicity and excessive reactive oxygen species (ROS) accumulation. Transcriptomic profiling revealed that Ba predominantly restored antioxidant pathways, notably reversing the Dex-induced downregulation of Sod3. Crucially, targeted functional assays demonstrated that SOD3 is an important mediator of this protection: Sod3 silencing abolished Ba's therapeutic efficacy, whereas Sod3 overexpression independently conferred profound resistance to oxidative injury.

Ba effectively ameliorates glucocorticoid-induced osteonecrosis by reconstituting the critical extracellular antioxidant barrier, a process closely associated with SOD3 upregulation. These findings establish Ba as a promising natural therapeutic agent and highlight extracellular redox modulation as a novel strategy for SONFH management.