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Ruxolitinib Delays Nucleus Pulposus Cell Senescence in Rat Intervertebral Discs
Honggang Hao1, Weidong Liang1, Shuwen Zhang2
1Department of Spine Surgery The First Affiliated Hospital of Xinjiang Medical University Urumqi China.
Background:
Intervertebral disc degeneration involves aging and senescence of nucleus pulposus cells (NPCs), and JAK/STAT signaling may contribute to this process. The aim of this study was to investigate the therapeutic effect of the JAK2 inhibitor, ruxolitinib, on NPC senescence.
Methods:
Control (third passage), Senescence (sixth passage), JAK inhibitor (ruxolitinib-treated), siRNA-NC (control siRNA-treated), and siRNA-JAK2 (JAK2-targeting siRNA-treated) groups of rat NPCs were established. Cell senescence ratios were determined by β-galactosidase staining and Edu staining was conducted to assess cell proliferation. Cell cycle and apoptosis were analyzed by flow cytometry and Aggrecan and Col II expression detected by immunofluorescence staining. Levels of IL-1β, IL-6, TNF-α, MMP-3, and MMP-13 were detected by ELISA, and p16, p21, p53, p-p53, JAK2, STAT3, p-JAK2, p-STAT3, ADAMTS4, and ADAMTS5 levels were examined by western blot.
Results:
More cell senescence was detected by β-galactosidase staining in the Senescence group than in the Control group, while cell proliferation was lower, apoptosis ratio higher, and the percentage of NPCs in G0/G1 phase higher. Levels of senescence-related proteins, including p16, p21, p53, and p-p53, were higher in the Senescence group than the Control group, as were those of IL-1β, IL-6, TNF-α, MMP-3, MMP-13, ADAMTS4, and ADAMTS5. Further, Aggrecan and Col II levels were lower in the Senescence group, while those of JAK2 and STAT3 (JAK2/STAT3 signaling pathway) were higher. Ruxolitinib reversed the changes described above to varying degrees, and the results were supported by those of experiments involving targeted silencing of JAK2.
Conclusions:
NPC senescence is characterized by low cell proliferation, a high apoptosis ratio, cell cycle arrest, and generation of senescence-associated secretory phenotypes. NPC senescence can be delayed by inhibiting JAK2/STAT3 signaling using ruxolitinib.
Insights
This study shows that inhibiting JAK2/STAT3 signaling with ruxolitinib can delay nucleus pulposus cell senescence, a key factor in intervertebral disc degeneration. This offers a potential therapeutic strategy for this condition.
Area of Science:
- Biomedical Science
- Cell Biology
- Molecular Biology
Background:
- Intervertebral disc degeneration is linked to aging and senescence of nucleus pulposus cells (NPCs).
- Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling pathways may play a role in NPC senescence.
Purpose of the Study:
- To investigate the therapeutic potential of ruxolitinib, a JAK2 inhibitor, in mitigating NPC senescence.
- To explore the role of JAK2/STAT3 signaling in the aging process of NPCs.
Main Methods:
- Established rat NPC cultures for control, senescence, ruxolitinib-treated, and JAK2-silenced groups.
- Assessed cell senescence, proliferation, cell cycle, apoptosis, and expression of key proteins and inflammatory markers using techniques like beta-galactosidase staining, Edu staining, flow cytometry, immunofluorescence, ELISA, and western blot.
Main Results:
- Senescent NPCs exhibited reduced proliferation, increased apoptosis, cell cycle arrest, and elevated levels of senescence markers, inflammatory cytokines, and matrix-degrading enzymes.
- The JAK2/STAT3 signaling pathway was upregulated in senescent NPCs, with increased levels of JAK2 and STAT3.
- Ruxolitinib treatment and JAK2 silencing partially reversed these senescence-associated changes.
Conclusions:
- NPC senescence is characterized by impaired proliferation, increased apoptosis, cell cycle arrest, and a senescence-associated secretory phenotype.
- Inhibiting the JAK2/STAT3 signaling pathway with ruxolitinib demonstrates a therapeutic effect in delaying NPC senescence.
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