UCHL3 drives ferroptosis in nucleus pulposus cells by deubiquitinating HMGB1 to activate NCOA4-mediated ferritinophagy.

Ferroptosis contributes to intervertebral disc degeneration (IVDD), yet its upstream regulatory mechanisms in nucleus pulposus cells (NPCs) remain unclear. Here, we identify a novel UCHL3-HMGB1-NCOA4 signaling axis that drives NPC ferroptosis. Transcriptomic and clinical analyses reveal that UCHL3 is upregulated in degenerative discs, with its expression correlating positively with both IVDD severity and ferroptosis markers. Mechanistically, UCHL3 deubiquitinates and stabilizes HMGB1 by cleaving Lys48-linked polyubiquitin chains, thereby promoting the accumulation of cytoplasmic HMGB1. This stabilized HMGB1 activates NCOA4-mediated ferritinophagy, initiating ferroptosis. Silencing UCHL3 or NCOA4 mitigates these effects. Furthermore, a polydopamine-based nanoplatform (PDA@si-UCHL3) achieves dual therapeutic action, silencing UCHL3 while scavenging ROS and chelating iron, which effectively inhibits ferroptosis and alleviates IVDD in rats. These findings establish UCHL3 as a pivotal regulator of NPC ferroptosis and highlight PDA@si-UCHL3 as a promising therapeutic strategy for IVDD.Abbreviations: AAV: adeno-associated virus; CCK-8: cell counting kit-8; Co-IP: co-immunoprecipitation; DUBs: deubiquitinating enzymes; ECM: extracellular matrix; GPX4: glutathione peroxidase 4; GSH: glutathione; HE: hematoxylin and eosin; HMGB1: high mobility group box 1; IF: immunofluorescence; IHC: immunohistochemistry; IP/MS: immunoprecipitation mass spectrometry; IVDD: intervertebral disc degeneration; IVDs: intervertebral discs; MRI: magnetic resonance imaging; NCOA4: nuclear receptor coactivator 4; NP: nucleus pulposus; NPCs: nucleus pulposus cells; PDA: polydopamine; PG: Pfirrmann grading; ROS: reactive oxygen species; siRNA: small interfering RNAs; SO-FG: Safranin O-Fast Green; TBHP: tert-butyl hydroperoxide; TEM: transmission electron microscope; UCHL3: ubiquitin C-terminal hydrolase L3; WT: wild-type.