Intervertebral disc (IVD) degeneration contributes largely to pathoanatomical and degenerative changes of spinal structure that increase the risk of low back pain. Apoptosis in nucleus pulposus (NP) can aggravate IVD degeneration, and increasing studies have shown that interventions targeting NP cell apoptosis can ameliorate IVD degeneration, exhibiting their potential for use as therapeutic strategies. Recent data have shown that advanced glycation end products (AGEs) accumulate in NP tissues in parallel with the progression of IVD degeneration and form a microenvironment of oxidative stress. This study examined whether AGEs accumulation aggravates NP cell apoptosis and IVD degeneration, and explored the mechanisms underlying these effects. We observed that the viability and proliferation of human NP cells were significantly suppressed by AGEs treatment, mainly due to apoptosis. Furthermore, activation of the mitochondrial apoptosis pathway was detected after AGEs treatment. In addition, the molecular data showed that AGEs could significantly aggravate the generation of mitochondrial reactive oxygen species and prolonged activation of the mitochondrial permeability transition pore, as well as the increased level of Bax protein and decreased level of Bcl-2 protein in mitochondria. These effects could be reduced by antioxidant (2-(2,2,6,6-Tetramethylpiperidin-1-oxyl-4-ylamino)-2-oxoethyl) triphenylphosphonium chloride (MitoTEMPO) and Visomitin (SKQ1). Importantly, we identified that impairment of Sirtuin3 (SIRT3) function and the mitochondrial antioxidant network were vital mechanisms in AGEs-induced oxidative stress and secondary human NP cell apoptosis. Finally, based on findings that nicotinamide mononucleotide (NMN) could restore SIRT3 function and rescue human NP cell apoptosis through adenosine monophosphate-activated protein kinase and peroxisome proliferator-activated receptor-γ coactivator 1α (AMPK-PGC-1α) pathway in vitro, we confirmed its protective effect on AGEs-induced IVD degeneration in vivo. In conclusion, our data demonstrate that SIRT3 protects against AGEs-induced human NP cell apoptosis and IVD degeneration. Targeting SIRT3 to improve mitochondrial redox homeostasis may represent a potential therapeutic strategy for attenuating AGEs-associated IVD degeneration.

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Although degeneration of the nucleus pulposus (NP) is a major contributor to intervertebral disc degeneration (IVDD) and low back pain, the underlying molecular complexity and cellular heterogeneity remain poorly understood. Here, a comprehensive single-cell resolution transcript landscape of human NP is reported. Six novel human NP cells (NPCs) populations are identified by their distinct molecular signatures. The potential functional differences among NPC subpopulations are analyzed. Predictive transcripts, transcriptional factors, and signal pathways with respect to degeneration grades are explored. It is reported that fibroNPCs is the subpopulation for end-stage degeneration. CD90+NPCs are observed to be progenitor cells in degenerative NP tissues. NP-infiltrating immune cells comprise a previously unrecognized diversity of cell types, including granulocytic myeloid-derived suppressor cells (G-MDSCs). Integrin 𝜶M (CD11b) and oxidized low density lipoprotein receptor 1 (OLR1) as surface markers of NP-derived G-MDSCs are uncovered. The G-MDSCs are found to be enriched in mildly degenerated (grade II and III) NP tissues compared to severely degenerated (grade IV and V) NP tissues. Their immunosuppressive function and alleviation effects on NPCs' matrix degradation are revealed in vitro. Collectively, this study reveals the NPC-type complexity and phenotypic characteristics in NP, thereby providing new insights and clues for IVDD treatment.