Abstract
BACKGROUND: Intervertebral disc degeneration (IDD) is a leading cause of spinal disorders, driven by oxidative stress-induced nucleus pulposus cell (NPC) apoptosis and extracellular matrix (ECM) degradation. Nuclear factor erythroid 2-related factor 2 (Nrf2) activators hold therapeutic promise due to their antioxidative properties. This study investigates the efficacy of RTA 408, a synthetic Nrf2-activating terpenoid, in mitigating oxidative damage and IDD progression. METHODS: In vitro, tert-butyl hydroperoxide (TBHP)-treated rat NPCs were pretreated with RTA 408 (10-100 nM) to assess antioxidative and antiapoptotic effects via CCK-8, ROS/DCFH-DA, MDA/SOD assays, Annexin V-FITC/PI staining, and mitochondrial membrane potential (JC-1) analysis. Western blotting evaluated Nrf2/ARE, NF-κB pathways, and ECM regulators (MMPs, ADAMTS5, collagen II, aggrecan). In vivo, a rat IDD model was established via coccygeal disc puncture, with RTA 408 (200/500 µg/kg, intraperitoneal) administered weekly. MRI, histopathology (H&E, Safranin O), and immunohistochemistry (aggrecan, MMP13, Nrf2) assessed disc degeneration over 4-8 weeks. RESULTS: In vitro, RTA 408 restored NPC viability, reduced ROS and MDA levels, and elevated SOD activity after TBHP exposure. It inhibited apoptosis (lower cleaved caspase-3 and BAX expression; higher BCL-2 levels) and mitochondrial depolarization. RTA 408 activated the Keap1/Nrf2/ARE pathway (promoted Nrf2 nuclear translocation and upregulated HO-1/NQO1) while suppressing NF-κB signaling (reduced phosphorylation of P65 and IκBα). ECM degradation was reversed (downregulated MMP3/9/13 and ADAMTS5; upregulated collagen II and aggrecan). In vivo, RTA 408 preserved disc structure, decreased Pfirrmann scores, and improved MRI indices (enhanced T2 signal intensity). Histopathological analysis confirmed reduced ECM loss and annulus fibrosus disruption, correlating with elevated Nrf2 expression and diminished MMP13 levels in nucleus pulposus. High-dose RTA 408 showed stronger therapeutic effects than low-dose treatment. CONCLUSIONS: RTA 408 mitigates oxidative stress-induced NPC apoptosis and ECM degradation via dual modulation of Nrf2/ARE activation and NF-κB suppression. Systemic administration of RTA 408 delays IDD progression in vivo, highlighting its therapeutic potential for degenerative spinal disorders. These findings support further clinical exploration of RTA 408 as a novel Nrf2-targeted therapy for IDD.