Ginsenoside-Rh2 Alleviates Pyroptosis of Nucleus Pulposus Cells by Activating Autophagy via HIF-1α to Retard the Progression of Intervertebral Disc Degeneration.

Macrophage infiltration and polarization have been increasingly observed in intervertebral disc (IVD) degeneration (IDD). However, their biological roles in IDD are still unrevealed. We harvested conditioned media (CM) derived from a spectrum of macrophages induced from THP-1 cells, and examined how they affect nucleus pulposus cells (NPCs) in vitro, by studying cell proliferation, extracellular matrix (ECM) synthesis, and pro-inflammation expression; and in vivo by injection CM in a rat IDD model. Then, high-throughput sequencing was used to detect differentially expressed genes (DEGs). Gene Ontology (GO), the Kyoto Encyclopedia of Genes and Genomes (KEGG), and protein-protein interaction (PPI) networks were used to further analysis. Higher CCR7+ (M1 marker) and CD206+ (M2 marker) cell counts were found in the degenerated human IVD tissues as compared with the control. Furthermore, the cell co-culture model showed M1CM attenuated NPC proliferation, downregulated the expression of ECM anabolic genes encoding aggrecan and collagen IIα1, upregulated the expression of ECM catabolic genes encoding MMP-13, and inflammation-related genes encoding IL-1β, IL-6, and IL-12, while M2CM showed contrasting trends. In IDD model, higher histological scores and lower disc height index were found following M1CM treatment, while M2CM exhibited opposite results. M1CM injection decreased ECM anabolic and increased ECM catabolic, as well as the upregulation of inflammation-related genes after 8 weeks treatment, while M2CM slowed down these trends. Finally, a total of 637 upregulated and 655 downregulated genes were detected in M1CM treated NPCs, and 975 upregulated genes and 930 downregulated genes in the M2CM groups. The top 30 GO terms were shown and the most significant KEGG pathway was cell cycle in both groups. Based on the PPI analysis, the five most significant hub genes were PLK1, KIF20A, RRM2, CDC20, and UBE2C in the M1CM groups and RRM2, CCNB1, CDC20, PLK1, and UBE2C in the M2CM groups. In conclusion, macrophage polarization exhibited diverse roles in IDD progression, with M1CM exacerbating cell proliferation suppression and IVD degeneration, while M2CM attenuated IDD development. These findings may facilitate the further elucidation of the role of macrophage polarization in IDD, and provide novel insights into the therapeutic potential of macrophages.

Danggui Sini decoction attenuates intervertebral disc degeneration by regulating ferroptosis in nucleus pulposus cells via the GJA1/cGAS/STING signaling axis.

Forsythiaside A attenuates intervertebral disc degeneration by suppressing PI3K/AKT/NF-κB-mediated NLRP3 inflammasome activation and pyroptosis.

PIM3 enhances nucleus pulposus cell function and extracellular matrix integrity via kinase-dependent activation of the Akt/mTOR signaling axis in intervertebral disc degeneration.

BackgroundThe method of action of Bushen Formula (BSHXF) in the treatment of intervertebral disc degeneration (IVDD) was uncovered in this work using in vivo and in vitro tests. To clarify the mechanism of action of BSHXF, we validated the rat intervertebral disc degeneration model and the nucleus pulposus cell degeneration model.MethodsIn an in vivo model of IVDD the study explores the impact of BSHXF on mitochondrial function, pro-inflammatory cytokines, pro-apoptotic factors, and matrix metalloproteinases. Additionally, it evaluates the induced degeneration of nucleus pulposus (NP) cells in an in vitro model stimulated by interleukin-1 β (IL-1β). The study measures the effects of BSHXF on both the inflammatory response and mitochondrial function.ResultsThe MRI results showed that BSHXF reduced intervertebral disc volume reduction and degradation of NP tissue. HE, SO-FG and immunofluorescence further confirmed the protective effect of BSHXF on degenerative intervertebral discs. BSHXF reduced the inflammatory levels of IL-6 IL-1β and TNF-α in degenerative intervertebral disc tissue. Meanwhile, JC-1, mPTP and ROS detection revealed that BSHXF can restore mitochondrial function by regulating the expression of antioxidant proteins, playing a protective role in NP cells. Finally, the WB results showed that BSHXF can alleviate IL-1β mediate the degeneration of NP cells. BSHXF can alleviate NP cell apoptosis by inhibiting the expression of bax, cleaved caspase-3, caspase-3, and cyt-c, and increasing the expression of Bcl-2.ConclusionThis study reveals that BSHXF inhibits the development of inflammatory factors, which may play a significant role in intervertebral disc degeneration. This implies that BSHXF is a suitable herbal medication for future research into inflammatory cytokine treatment.

Intervertebral disc degeneration (IDD) is the main pathological factor resulting in low back pain (LBP), the leading cause of disability globally. Inflammatory response and extracellular matrix (ECM) degradation are critical pathological features in the development of IDD. Gastrodin (GAS), a phenol compound isolated from Gastrodia elata Blume, plays an anti-inflammatory role in experimental models of multiple human diseases. Our study aimed to elucidate whether GAS alleviates TNF-α-induced inflammation in nucleus pulposus (NP) cells and IDD in vivo. The cytotoxicity of GAS was assessed by CCK-8 assay. Rat primary NP cells were stimulated with TNF-α to induce inflammatory response. The expression of proinflammatory cytokines, catabolic genes, and anabolic genes was detected by RT-qPCR, western blotting, and immunofluorescence staining. NF-κB and MAPK pathway activation was determined through western blotting and immunofluorescence staining. The IDD rat model was established by using percutaneous needle puncture. The therapeutic effects of GAS were confirmed by histology analysis. We found that TNF-α stimulation enhanced proinflammatory cytokine (COX2, iNOS, IL-6, and IL-1β) expression in NP cells, which was reversed by GAS treatment. GAS offset TNF-α-induced upregulation in catabolic gene (MMP3, MMP9, and MMP13) expression and downregulation in anabolic gene (Collagen II, SOX9, and Aggrecan) expression. The loss of ECM in TNF-α-treated NP cells was mitigated by GAS treatment. Mechanically, GAS abolished TNF-α-induced increase in p-IKKα, p-IKKβ, p-IκBα, p-p65, p-ERK, p-p38, and p-JNK protein levels in NP cells. In puncture-induced IDD rat models, GAS administration improved intervertebral disc (IVD) structure, increased Collagen II expression, and reduced the levels of proinflammatory factors in IVDs. Overall, GAS alleviates the inflammation and ECM degradation in NP cells via inhibiting NF-κB and MAPK pathway activation and alleviates IDD in vivo, which may be a novel treatment strategy for IDD.

The imbalance between TIMP3 and matrix-degrading enzymes plays an important role in intervertebral disc degeneration

The role of the miR-4306/PAK6 axis in degenerative nucleus pulposus cells in human intervertebral disc degeneration

Bardoxolone methyl breaks the vicious cycle between M1 macrophages and senescent nucleus pulposus cells through the Nrf2/STING/NF-κB pathway

Intervertebral disc degeneration (IDD) is a leading cause of global disability. This study aimed to systematically investigate the role of SUMOylation in the pathogenesis of IDD through integrated transcriptomic analysis. Bioinformatics analysis was performed on public transcriptomic datasets (GEO). Human nucleus pulposus (NP) cells were used for experimental validation, followed by RNA sequencing and mechanistic studies. A rat IDD model was established to assess the effects of modulating SUMOylation in vivo. SUMOylation was found to be significantly involved in IDD. The expression of the deSUMOylating enzyme SENP1 was downregulated in degenerated NP cells. Overexpression of SENP1 alleviated cellular senescence and extracellular matrix metabolic dysfunction. Further mechanistic studies revealed that SENP1 exerted its protective effects by inhibiting both apoptosis and pyroptosis in NP cells. In the rat IDD model, intervention targeting SUMOylation markedly slowed disease progression. This study demonstrates that SENP1 attenuates IDD progression by suppressing apoptosis and pyroptosis, highlighting the central role of SUMOylation in IDD pathogenesis and identifying the SENP1 axis as a potential therapeutic target for this debilitating condition.

Nucleus pulposus (NP) cell apoptosis and extracellular matrix (ECM) degradation constitute the two major pathological hallmarks of intervertebral disc degeneration (IVDD). Inhibiting these deleterious processes represents an effective strategy for attenuating IVDD progression. Sirtuin 7 (SIRT7), a member of the sirtuin family, plays a critical role in modulating gene expression, mediating cellular stress adaptation, and facilitating DNA repair. While SIRT7 has demonstrated therapeutic potential across diverse pathological contexts, its specific contribution to IVDD pathogenesis remains elusive. This study aimed to delineate the functional contribution of SIRT7 to IVDD progression and unravel its molecular mechanisms. We quantified SIRT7 levels by immunohistochemistry (IHC) in degenerative human and rat NP tissues, and by quantitative reverse transcription polymerase chain reaction (qRT-PCR) in tert-butyl hydroperoxide (TBHP)-treated NP cells. To evaluate the protective capacity of SIRT7 overexpression, we conducted multifaceted analyses encompassing oxidative stress markers, apoptotic indices, ECM turnover, and nuclear factor kappa B (NF-κB) cascade activity in TBHP-challenged NP cells, utilizing reactive oxygen species detection probes, mitochondrial membrane potential indicators, Hoechst 33342 nuclear staining, qRT-PCR, western blotting, and immunofluorescence techniques. Its therapeutic potential was subsequently validated through magnetic resonance imaging and comprehensive histopathological evaluation (hematoxylin and eosin, Safranin O/Fast Green, and Masson trichrome staining) combined with IHC in a rat IVDD puncture model. SIRT7 was consistently downregulated in degenerated human and rat NP tissues as well as TBHP-treated NP cells, concomitant with elevated NF-κB pathway activation. SIRT7 overexpression in TBHP-stimulated NP cells effectively attenuated oxidative stress, apoptosis, and ECM degradation. Mechanistically, SIRT7 overexpression may exert inhibitory effects on NF-κB signaling. Consistently, SIRT7 overexpression in the rat IVDD model decreased NF-κB activity, reduced NP cell apoptosis and ECM depletion, eventually ameliorating disc degeneration. Our findings demonstrate that SIRT7 expression declines progressively during IVDD development. SIRT7 overexpression protects against NP cell apoptosis and ECM degradation, and this protective effect correlates with inhibition of the NF-κB pathway. These findings suggest tha SIRT7 is a guardian of NP homeostasis and highlight its substantial promise as a molecular target for IVDD therapeutics.

The destruction of the low oxygen microenvironment in nucleus pulposus (NP) cells played a critical role in the pathogenesis of intervertebral disc degeneration (IVDD). The purpose of this study was to determine the potential role of integrin alpha 6 (ITG α6) in NP cells in response to high oxygen tension (HOT) in IVDD. Immunofluorescence staining and western blot analysis showed that the levels of ITG α6 expression were increased in the NP tissue from IVDD patients and the IVDD rat model with mild degeneration, which were reduced as the degree of degeneration increases in severity. In NP cells, the treatment of HOT resulted in upregulation of ITG α6 expression, which could be alleviated by blocking the PI3K/AKT signaling pathway. Further studies found that ITG α6 could protect NP cells against HOT-induced apoptosis and oxidative stress and protect NP cells from HOT-inhibited ECM protein synthesis. Upregulation of ITG α6 expression by HOT contributed to maintaining NP tissue homeostasis through the interaction with hypoxia-inducible factor-1α (HIF-1α). Furthermore, silencing of ITG α6 in vivo could obviously accelerate puncture-induced IVDD. Taken together, these results revealed that the increase of ITG α6 expression by HOT in NP cells might be a protective factor in IVD degeneration as well as restore NP cell function.

Intervertebral disc degeneration (IDD) is integral in lower back pain and involves complex pathophysiological processes, including nucleus pulposus (NP) cell apoptosis and extracellular matrix (ECM) breakdown. Palmatine (PLT), an isoquinoline alkaloid extracted from Fibraurea recisa Pierre of the family Menispermaceae, is recognised for its anti-inflammatory, antioxidant, and neuroprotective effects. Nevertheless, researches have not well explored the impact of PLT on IDD. This investigation aimed at determining the impact of PLT on oxidative stress caused by tert‑butyl hydroperoxide (TBHP) and exploring its potential as a therapeutic agent and its mechanisms in IDD. Potential anti-IDD targets of PLT were identified using network pharmacology and bioinformatics methods and evaluated using Gene Ontology analysis. The method of molecular docking helped elucidate the interaction mode and connections between PLT and transcription factor EB (TFEB). Cellular thermal shift assays and cycloheximide chase experiments confirmed direct interactions between PLT and TFEB. NP cell apoptosis, ECM levels, endoplasmic reticulum stress (ERS), autophagy, and TFEB expression were evaluated using western blotting, TUNEL staining, EdU staining, flow cytometry, immunofluorescence, and alcian blue staining. Functional IDD recovery was evaluated using MRI and X-ray, haematoxylin-eosin (HE) staining, safranin O/fast green staining, and immunohistochemical (IHC) staining. Moreover, needle puncture was used to establish an in vivo rat model of IDD to examine the therapeutic efficacy of PLT. PLT markedly mitigated ERS and inhibited TBHP-induced ECM degradation and NP cell apoptosis by activating TFEB and upregulating autophagy. In the IDD rat model, PLT improved annulus fibrosus (AF) and NP morphology and structure. These findings demonstrate that PLT alleviates IDD progression by upregulating TFEB; therefore, TFEB represents a potential novel therapeutic target. Moreover, this study reveals for the first time that PLT inhibits ERS by enhancing TFEB-mediated autophagy, thereby reducing NP cell apoptosis and ECM degradation, thus providing valuable insights into the key pharmacological mechanisms of PLT.

Intervertebral Disc degeneration (IDD) is one of the leading causes of disability, and current therapies are ineffective. Phosphodiesterase 4B (PDE4B) plays an essential role in regulating the activation of nuclear factor E2-related factor 2 (Nrf2), while Nrf2 regulates ferroptosis. However, it is still unknown whether PDE4B is involved in the development of IDD. In this study, we explored the role of PDE4B on ferroptosis and Nrf2 in IDD pathogenesis by in vivo and in vitro experiments. The findings suggested that the expressions of PDE4B, ASCL4, and TRFC were significantly upregulated, and the expression of Nrf2 was significantly downregulated in nucleus pulposus (NP) tissues from human IDD patients dependent on IDD degeneration. Overexpression of PDE4B (PDE4B-OE) in NP cells upregulated the expression of ASCL4 and TRFC, and downregulated the expression of Nrf2. Meanwhile, the level of cytokine and oxidative stress were upregulated. Ferroptosis inhibitor Fer-1 or Nrf2 activator dimethyl fumarate (DMF) suppressed the effect of PDE4B-OE, while ferroptosis inducer elastin enhanced the effect of PDE4B-OE. In the IDD rat model, PDE4 inhibitor roflumilast, ferroptosis inhibitor Fer-1, or Nrf2 activator dimethyl fumarate (DMF) delayed IDD pathogenesis. While administration of ferroptosis inducer elastin enhanced IDD pathogenesis. Combination with PDE4B inhibitor and ferroptosis inhibitor Fer-1 significantly synergistic reversed IDD pathogenesis. While combination with PDE4B inhibitor or Nrf2 activator and elastin also decreased the degree of the IDD. The IHC suggested PDE4 inhibitor downregulated the expression of ASCL4 and TRFC. However, the combination effect of the Nrf2 activator was not obvious. Our study suggested that aberrant PDE4B activation in NP tissues induces pathological changes in IDD mediated by ferroptosis, and PDE4 inhibitor reveres the process of IDD by suppressing ferroptosis, and has a synergic effect with ferroptosis inhibitor. So PDE4B inhibition may be a potential therapeutic strategy for IDD.

Lower back pain (LBP) is a worldwide health problem associated with significant economic and social burden. Intervertebral disc degeneration (IVDD) is a leading cause of LBP. Several studies show that the death of nucleus pulposus cells (NPCs), abnormal metabolism of the extracellular matrix (ECM), and inflammatory response are the key mechanisms behind the pathogenesis of IVDD. Circular RNAs (circRNAs) are key regulators of gene expression and play a significant role in regulating NPCs death, ECM homeostasis, and inflammatory response by acting as microRNAs (miRNAs) sponges in IVDD. However, the regulatory role of circRNAs in mediating IVDD remains unknown. This review comprehensively describes the normal anatomic structure and function of IVD, the pathogenesis of IVDD, the characteristics, synthesis, mechanisms, and function of circRNAs. Moreover, we highlighted the 23 circRNAs that mediate ECM metabolism, 16 circRNAs that mediate NPCs apoptosis, circ_0004354 and circ_0040039 that mediate NPCs pyroptosis, and 5 circRNAs that mediate inflammatory response in IVDD. In addition, this review presents suggestions for future studies, such as the need for further investigation on ferroptosis-related circRNAs in IVDD. This review could provide novel insights into the pathogenesis and treatment of IVDD.