FOXO3 counteracts oxidative stress-induced nucleus pulposus cell senescence via BNIP3-mediated mitophagy in intervertebral disc degeneration.

Oxidative stress and mitochondrial reactive oxygen species (ROS) accumulation are central drivers of nucleus pulposus (NP) cell senescence and intervertebral disc degeneration (IVDD), yet the transcriptional programs that maintain mitochondrial redox homeostasis in NP cells remain poorly defined. In this study, we investigated the role of forkhead box O3 (FOXO3) in regulating NP cell senescence and mitophagy during IVDD. We found that FOXO3 expression was significantly reduced in degenerated disc tissues and in senescent NP cells. In vitro, FOXO3 overexpression markedly alleviated H2O2-induced senescence, as shown by reduced SA-β-gal positivity, downregulation of p16 and senescence-associated catabolic factors, and partial restoration of extracellular matrix-related proteins. Transcriptomic profiling revealed that FOXO3 suppressed senescence-, NF-κB-, and matrix degradation-associated pathways while activating autophagy-related programs. Mechanistically, FOXO3 enhanced autophagic flux and promoted BNIP3-dependent mitophagy, accompanied by improved mitochondrial ultrastructure, preservation of mitochondrial membrane potential, and reduced mitochondrial reactive oxygen species accumulation. Importantly, BNIP3 knockdown partially abolished the anti-senescent effects of FOXO3, supporting BNIP3-mediated mitophagy as a key downstream mechanism. In vivo, adeno-associated virus-mediated FOXO3 overexpression attenuated puncture-induced disc degeneration, reduced p16 and MMP13 expression, and restored aggrecan and collagen II levels. Collectively, these findings identify FOXO3 as a critical regulator of mitochondrial quality control and NP cell senescence, and suggest that targeting the FOXO3-BNIP3-mitophagy axis may represent a promising therapeutic strategy for IVDD.