Inhibition of the JAK2/STAT3 Pathway Attenuates D-Galactose-Induced Nucleus Pulposus Cell Senescence and Intervertebral Disc Degeneration.

OBJECTIVE: This study aimed to investigate the effect of Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) pathway inhibition on D-galactose (D-gal)-induced senescence in nucleus pulposus cells (NPCs) and its potential to delay intervertebral disc degeneration (IVDD), as well as to investigate the underlying mechanisms. METHODS: A cellular senescence model was established by treating rat NPCs with D-gal. The model was then intervened with a JAK2/STAT3 pathway inhibitor (ruxolitinib) or JAK2-specific small interfering RNA (siRNA). Cellular senescence was evaluated by senescence-associated β-galactosidase (SA-β-gal) staining. The expression of senescence markers (p16, p21, and p53), extracellular matrix (ECM) components (aggrecan and collagen II), catabolic enzymes (ADAMTS-4, ADAMTS-5, MMP-3, and MMP-13), and JAK2/STAT3 pathway proteins was analyzed by western blotting and immunofluorescence. The levels of inflammatory factors (interleukin [IL]-1β, IL-6, and tumor necrosis factor-α [TNF-α]) and advanced glycation end-products (AGEs) were measured by enzyme-linked immunosorbent assay (ELISA). Cell proliferation, apoptosis, and cell cycle distribution were assessed using cell counting kit-8 (CCK-8) and flow cytometry. In a parallel in vivo study, a rat model of IVDD was induced by D-gal and treated with the JAK inhibitor. Disc degeneration was evaluated by magnetic resonance imaging (MRI) and histopathological examination after 8 weeks. RESULTS: Both in vitro and in vivo, inhibition of the JAK2/STAT3 pathway, either pharmacologically or genetically, effectively attenuated D-gal-induced effects. It suppressed the phosphorylation of STAT3, reduced the expression of SA proteins (p16, p21, and p53), ECM catabolic enzymes (ADAMTS-4, ADAMTS-5, MMP-3, and MMP-13), and proinflammatory cytokines (IL-1β and IL-6). Consequently, this inhibition decreased SA-β-gal positivity, alleviated cell cycle arrest and apoptosis, and enhanced the synthesis of aggrecan and collagen II in NPCs. In the rat model, JAK inhibitor treatment improved MRI scores, restored disc signal intensity, and ameliorated histopathological degeneration. CONCLUSION: : Inhibition of the JAK2/STAT3 pathway reduced the expression of inflammatory factors and oxidative stress markers in D-gal-treated NPCs. It also suppressed ECM degradation and apoptosis, delayed cellular senescence, and attenuated the progression of IVDD in rats.