Pterostilbene mitigates intervertebral disc degeneration via dual regulation of mitophagy and NLRP3 ubiquitination to inhibit nucleus pulposus cell pyroptosis.

BACKGROUND

Intervertebral disc degeneration (IVDD) is a major contributor to low back pain and is characterized by mitochondrial dysfunction, inflammation, and regulated cell death in nucleus pulposus (NP) cells. NLRP3 inflammasome-mediated pyroptosis plays a pivotal role in disc degeneration, whereas mitophagy limits mitochondrial damage and inflammasome activation. Emerging evidence indicates that pterostilbene (PTE) exhibits diverse pharmacological activities; however, its role in attenuating IVDD remains insufficiently understood.

OBJECTIVE

To investigate whether PTE attenuates IVDD by regulating mitophagy and NLRP3 inflammasome-mediated pyroptosis, and to elucidate the underlying molecular mechanisms.

STUDY

DESIGN

This study combined network pharmacology analysis, in vitro cellular experiments, and an in vivo rat IVDD model to evaluate the therapeutic effects and mechanisms of PTE.

METHODS

Network pharmacology was used to predict the cytoprotective potential of PTE. In vitro, LPS-stimulated NP cells were employed to evaluate inflammation, extracellular matrix degradation, pyroptosis, mitochondrial function, and mitophagic flux. Pharmacological inhibitors were used to interrogate mitophagy-dependent mechanisms, while Western blotting, co-immunoprecipitation, and related assays were performed to assess signaling pathways, protein stability, and ubiquitination. In vivo, a rat IVDD model was established to evaluate the therapeutic effects of PTE through radiological and histopathological analyses.

RESULTS

PTE attenuated LPS-induced inflammatory responses and extracellular matrix degradation in NP cells. It suppressed NLRP3 inflammasome-mediated pyroptosis and restored mitochondrial function by enhancing mitophagic flux, whereas inhibition of mitophagy partially reversed these protective effects. Mechanistically, PTE activated the AMPK/mTOR/ULK1 signaling pathway to promote mitophagy. In parallel, PTE facilitated ubiquitination and proteasomal degradation of NLRP3, predominantly through K48-linked polyubiquitination, while K63-linked modification may contribute to regulatory signaling. Notably, this ubiquitination-mediated degradation occurred independently of mitophagy. In vivo, PTE significantly alleviated disc degeneration, reduced NLRP3, caspase-1, and p-mTOR expression, and increased collagen II, SOX9, PINK1, LC3, and p-AMPK levels.

CONCLUSION

PTE attenuates IVDD progression by coordinately enhancing mitophagy and promoting K48-linked ubiquitin-proteasome-dependent degradation of NLRP3. These two mechanisms operate independently yet synergistically to suppress inflammasome activation, providing mechanistic insight into PTE as a potential disease-modifying therapeutic strategy for IVDD.