Search Results (115)

Current therapeutic strategies for intervertebral disc degeneration (IDD)-related low back pain are limited to symptomatic alleviation. Notochordal cells (NCs), as progenitor cells of the nucleus pulposus (NP), lead us to develop innovative NC-based new therapies for IDD. A total of 40 NP specimens, obtained according to IDD criteria with defined Pfirrmann grades and histological degeneration score, were categorized as either normal (Grade II) or degenerated (Grade IV). An IDD model was established in SD rats by needle puncture of the annulus fibrosus. Degenerated NP tissue was identified using MRI, H&E, Safranin O, and Masson staining. NCs and NP cells (NPCs) were isolated and identified based on specific cellular markers. Furthermore, mRNA-seq was performed to profile gene expression in these cells. GO annotation and KEGG pathway analysis were employed to perform functional enrichment analysis of the differentially expressed genes (DEGs). Cell viability was assessed using the CCK-8 assay. An in vitro cell degeneration model was established by treating NPCs with TBHP. Analysis of specific marker expression was performed using Western blotting, immunohistochemistry, and immunofluorescence. We found that the number of NCs in degenerated NP tissues was significantly reduced compared to those in normal NP tissues, but a small amount of notochordal cell markers could still be detected. Analysis of sequencing data identified 2391 upregulated and 3813 downregulated DEGs. GO enrichment analysis indicated that these DEGs were significantly associated with regulatory signals including cellular senescence and oxidative stress. KEGG pathway analysis further revealed that the DEGs were primarily enriched in the TNF and HIF-1 signaling pathways. Subsequent screening identified the top 10 key genes potentially related to IDD: Sod2, Cxcl12, Spp1, Fn1, Cat, Il6, Ccl2, Igf1, Fgf2, and Acta2. Collectively, our findings establish a clear link between SOD2/CAT and the pathogenesis of IDD. SOD2 and CAT may serve as promising new potential therapeutic targets for IDD by inhibiting oxidative stress and cellular senescence in NPCs. Full article

Obesity is a recognized risk factor for intervertebral disc (IVD) degeneration, a condition characterized by the progressive loss of extracellular matrix components in the nucleus pulposus. Elevated circulating leptin levels in obese individuals contribute to this degeneration by upregulating matrix metalloproteinase-1 (MMP-1) expression. Targeting MMP-1 expression with low-toxicity natural compounds may provide a promising strategy to prevent or mitigate IVD degeneration. In this study, we examined the effects of cinnamaldehyde (CA), a natural compound derived from Cinnamomum osmophloeum Kaneh, on leptin-induced MMP-1 expression in human IVD cartilage endplate-derived stem cells (SV40 cell line). Our results showed that leptin induced MMP-1 expression via activation of leptin receptor-mediated JAK2/STAT3, JAK2/RhoA/STAT3, and RhoA/ERK1/2/NF-κB signaling pathways. CA significantly reduced MMP-1 expression by inhibiting phosphorylation of the leptin receptor and STAT3 and blocking RhoA and NF-κB activation, without affecting JAK2 and ERK1/2 phosphorylation. These findings suggest that CA suppresses leptin-induced MMP-1 expression by modulating specific signaling pathways, highlighting its potential as a therapeutic agent for IVD degeneration associated with obesity. Full article

Background: Intervertebral disc degeneration (IVDD) is closely linked to low back pain (LBP), a leading cause of disability worldwide. IVDD is characterized by the loss of proteoglycans (PGs), extracellular matrix (ECM) degradation, and reduced hydration of the nucleus pulposus (NP). Extracellular vesicles (EVs) derived from human umbilical cord mesenchymal stem cells (hUC-MSCs) exhibit tissue repair and immunomodulatory effects and are emerging as promising cell-free therapeutics. Methods: We established a rat IVDD model via fluoroscopy-guided needle puncture of three consecutive coccygeal discs and confirmed degeneration through Alcian Blue and hematoxylin & eosin (H&E) staining. The gene expression of inflammatory and pain markers (ADRβ2, COMP, CXCL1, COX2, PPTA, MMP13, YKL40) was measured by qPCR. Subsequently, we implanted hUC-MSCs or EVs to evaluate their reparative potential. Results: Upregulation of inflammatory and pain genes in IVDD was associated with an immunomodulatory response. Tracking DiI-labelled hUC-MSCs and EVs revealed enhanced survival of hUC-MSCs, retention of EVs, and dispersion within rat tail discs; EVs showed greater retention than hUC-MSCs. Implanted EVs were internalized by NP cells and remained within degenerative IVDs. EVs passively diffused, accumulated at the injury site, interacted with host cells, and enhanced function, as shown by increased expression of human chondrocyte-related markers (SOX9, TGFβ1, TGFβ2, COL2) compared to hUC-MSC treatment. Histological analysis of two weeks post-transplantation showed NP cellular patterns resembling chondromas in treated discs. EVs integrated into and distributed within degenerated NP regions, with greater glycosaminoglycan (GAG) content. Conclusions: Overall, hUC-MSC EVs demonstrated superior regenerative capacity, supporting a safe, cell-free strategy for disc repair. Full article

Intervertebral disc degeneration is a leading cause of back and leg pain and a major contributor to disability worldwide. Despite its prevalence, treatments remain limited due to incomplete understanding of its pathology. In vivo models pose challenges for controlled conditions, while in vitro cell cultures lack key cell–cell and cell–matrix interactions. To address these limitations, we developed a novel tissue slice culture model of mouse discs, in which intact mouse discs were sliced down to 300 μm thickness with a vibratome and cultured ex vivo at various time points. The cell viability, matrix components, structure integrity, inflammatory responses, and macrophage interactions were evaluated with biochemistry, gene expression, histology, and 3D imaging analyses. Disc slices maintained structural integrity and cell viability, with preserved extracellular matrix in the annulus fibrosus (AF) and mild degeneration in nucleus pulposus (NP) by day 5. Interleukin-1 (IL-1) induced disc degeneration manifested by increased glycosaminoglycan release in media and reduced aggrecan and collagen II mRNA levels in disc cells. Cultured disc slices promoted macrophages towards pro-inflammatory phenotype with elevated mRNA levels of il-1α, il-6, and inos. Macrophage overlay and 3D imaging demonstrated macrophage infiltration into the NP and AF tissues up to ~100 µm in depth. The disc tissue slice model captures key features of intervertebral discs and can be used for investigating mechanisms of disc degeneration and therapeutic evaluation. Full article

Doxorubicin (DOX) is widely used for the treatment of several tumors, but considerable dose-dependent side effects on many normal tissues, including bones, have been reported. The aim of the present study was to follow for the first time the kinetics of DOX accumulation/clearance in the non-vascularized intervertebral disc (IVD), as well as to assess the drug’s biological action in the annulus fibrosus (AF) and nucleus pulposus (NP) IVD cells and tissues. DOX was administered intravenously to rabbits before the isolation of IVDs, in which DOX quantification was performed using a highly sensitive LC-HRMS/MS analytical method. The effect of the drug on IVD cells’ physiology was assessed in vitro, while IVD tissue quality post-DOX administration was studied in vivo through histological analysis. DOX delivery was found significantly lower in the IVD compared to the highly vascularized skin, declining from the outer AF to the inner NP. The low DOX concentrations reaching the IVDs had marginal effects on cells’ viability, intracellular redox status, and p38 MAPK activation, while they did not evoke cellular senescence. Most importantly, the drug did not negatively affect ECM integrity, as collagen and proteoglycan content remained stable in vitro and in vivo. Full article

The intervertebral disc, an anatomical compartment interposed between vertebral bodies, plays a key role in spine flexibility and compression loading. It comprises three tissues: the nucleus pulposus, the annulus fibrosus, and the end plates. Degeneration-related changes in the extracellular matrix of the nucleus pulposus upon ageing or pathological conditions prompted the present investigation into the impact of proteoglycan reduction, the main constituent of the healthy nucleus pulposus, on its physicochemical properties and cellular phenotypical changes. To mimic the native extracellular matrix, three-dimensional NP-mimicking constructs were developed using a biomimetic hydrogel composed of collagen type I, collagen type II, and proteoglycans. This system was fabricated using a bottom-up approach, employing highly pure monomeric collagen types I and II, which were induced to form a reconstituted fibrillar structure closely resembling the natural NP microenvironment. A comprehensive physicochemical characterization was conducted at varying proteoglycan percentages using scanning electron microscopy (SEM), FTIR, rheological tests, and water retention property analysis. The effect of microenvironment changes on the phenotype of nucleus pulposus cells was studied by their encapsulation within the various collagen–proteoglycan hydrogels. The morphological and immunochemistry analysis of the cells was performed to study the cell–matrix adhesion pathways and the expression of the cellular regulator hypoxia-inducible factor 1 alpha. These were linked to the analysis of the synthesis of healthy or pathological extracellular matrix components. The findings reveal that the reduction in proteoglycan content in the nucleus pulposus tissue triggers a pathological pathway, impairing the rheological and water retention properties. Consequently, the cell phenotypes are altered, inducing the synthesis of collagen type I rather than securing the natural physiological remodelling process by the synthesis of collagen type II and proteoglycans. Identifying the proteoglycan content threshold that triggers these pathological phenotypical changes could provide new diagnostic markers and early therapeutic strategies for intervertebral disc degeneration. Full article

Apoptosis plays a crucial role in the progression of intervertebral disc degeneration (IVDD), a significant cause of chronic low back pain. This review explores disc cell apoptosis’s cellular and molecular mechanisms, focusing on nucleus pulposus, annulus fibrosus, and cartilage endplates cells. Apoptotic pathways—intrinsic (mitochondrial), extrinsic (death receptor-mediated), ER stress-mediated, and autophagy-related—are activated by oxidative stress, inflammation, mechanical load, and metabolic disturbances like hyperglycemia. Diabetes exacerbates disc cell apoptosis through AGE-RAGE signaling and mitochondrial dysfunction. Inflammation further amplifies apoptotic cascades via cytokine signaling and ROS generation. The review also examines emerging therapeutic strategies, including antioxidants (e.g., MitoQ, resveratrol), anti-inflammatory agents (e.g., cytokine inhibitors), autophagy modulators (e.g., rapamycin, metformin), and stem cell and gene therapies. While promising preclinical results exist, challenges such as poor bioavailability and clinical translation remain. Enhanced understanding of apoptosis pathways informs future cellular preservation and matrix integrity treatments. Based on a comprehensive literature search from 2000 to 2025, this narrative review synthesizes current knowledge, identifies knowledge gaps, and discusses translational potential. Our findings support a paradigm shift toward mechanism-based therapies that address the root cause of IVDD rather than symptomatic relief alone. Full article

Background: Intervertebral disc degeneration (IVDD) is a major cause of chronic back pain. Recent studies suggest that ferroptosis, a form of cell death, contributes to the degeneration of nucleus pulposus cells (NPCs). This study explores a novel therapeutic strategy using cell-free fat extract (CEFFE), rich in cytokines, to mitigate IVDD by inhibiting oxidative stress-induced ferroptosis. Methods: In vitro, NPC degeneration was induced by TNF-α/TBHP. The effects of CEFFE on matrix metabolism were evaluated using Western blotting, RT-qPCR, and high-density culture, with regenerative effects measured via CCK-8 assays. Ferroptosis was assessed by Western blotting, immunofluorescence, and electron microscopy. In vivo, rats with caudal IVDD were treated with CEFFE for 4 weeks, and therapeutic efficacy was evaluated through imaging and histological analysis. Results: In vitro, CEFFE reduced TNF-α-induced inflammation and promoted matrix synthesis by inhibiting MAPK and NF-κB pathways. It also activated NRF2 to prevent TBHP-induced ferroptosis. In rats, CEFFE facilitated nucleus pulposus repair and significantly slowed disc degeneration. Conclusions: CEFFE is a promising strategy to delay IVDD progression by inhibiting ferroptosis, offering potential therapeutic benefits for disc degeneration. Full article

Intervertebral disc (IVD) disease is typically characterized by the degradation of IVD tissue, secretion of inflammatory and painful factors, and hyperinnervation of the disc. The pro-inflammatory cytokine interleukin-1β (IL-1β) has been regarded as a principal factor in orchestrating disc degeneration. Link N (LN) is a peptide derived from the link protein that has been shown to promote extracellular disc regeneration even in an inflammatory milieu; however, no mechanism(s) has been described for their behaviour to date. Building on prior studies on LN, we hypothesize that LN directly inhibits IL-1β. IVD degeneration was experimentally induced in New Zealand white rabbits, followed by the injection of either sLN or saline as the vehicle control. To determine the expression of markers of pain, histology was performed. Cultured human Nucleus Pulposus disc cells (hNP) were used to determine the effects of LN on IL-1β-induced changes in gene expression, including the effects on IL-1β, TNFα, and IL6 signalling. Isolated murine dorsal root ganglia (DRG) neurons were used to assess the effect of LN on IL-1β-induced neuronal hyperactivity. LN significantly reduced IL-1β-induced NF-κB activation in a dose-dependent manner in disc cells and was further able to modulate IL-1β-induced gene expression, inflammatory mediators, and neurotrophic factors. Peptide docking simulations revealed that LN could interact with IL-1β. A direct interaction of LN and IL-1β was revealed through co-immunoprecipitation experiments. Although IL-1β was able to hypersensitize DRG neurons following a seven-day exposure, as demonstrated by Ca²⁺ imaging, this effect was significantly blunted when co-treated with LN. LN demonstrates a novel mechanism of action by directly inhibiting IL-1β, in addition to mitigating IL-1β-induced hypersensitivity in DRG neurons. These data suggest a potential role for LN in reducing discogenic pain. Full article

The mammalian target of rapamycin (mTOR), a serine/threonine kinase, promotes cell growth and inhibits autophagy. The following two complexes contain mTOR: mTORC1 with the regulatory associated protein of mTOR (RAPTOR) and mTORC2 with the rapamycin-insensitive companion of mTOR (RICTOR). The phosphatidylinositol 3-kinase (PI3K)/Akt/mTOR signaling pathway is important in the intervertebral disk, which is the largest avascular, hypoxic, low-nutrient organ in the body. To examine gene-silencing therapeutic approaches targeting PI3K/Akt/mTOR signaling in degenerative disk cells, an in vitro comparative study was designed between small interfering RNA (siRNA)-mediated RNA interference (RNAi) and clustered regularly interspaced short palindromic repeat (CRISPR)–CRISPR-associated protein 9 (Cas9) gene editing. Surgically obtained human disk nucleus pulposus cells were transfected with a siRNA or CRISPR–Cas9 plasmid targeting mTOR, RAPTOR, or RICTOR. Both of the approaches specifically suppressed target protein expression; however, the 24-h transfection efficiency differed by 53.8–60.3% for RNAi and 88.1–89.3% for CRISPR–Cas9 (p < 0.0001). Targeting mTOR, RAPTOR, and RICTOR all induced autophagy and inhibited apoptosis, senescence, pyroptosis, and matrix catabolism, with the most prominent effects observed with RAPTOR CRISPR–Cas9. In the time-course analysis, the 168-h suppression ratio of RAPTOR protein expression was 83.2% by CRISPR–Cas9 but only 8.8% by RNAi. While RNAi facilitates transient gene knockdown, CRISPR–Cas9 provides extensive gene knockout. Our findings suggest that RAPTOR/mTORC1 is a potential therapeutic target for degenerative disk disease. Full article

Intervertebral disc degeneration is a leading cause of chronic low back pain, affecting millions globally. Regenerative medicine, particularly cell-based therapies, presents a promising therapeutic strategy. This study evaluates the comparative efficacy of two biomaterials—hyaluronic acid (HA) and alginate—as carriers for nucleus pulposus (NP) cell transplantation in a beagle model of induced disc degeneration. NP cells were isolated, cultured, and injected with either HA or alginate into degenerated discs, with saline and non-cell-loaded carriers used as controls. Disc height index, T2-weighted MRI, and histological analyses were conducted over a 12-week follow-up period to assess reparative outcomes. Imaging revealed that both carrier and cell-loaded treatments improved outcomes compared to degenerative controls, with cell-loaded carriers consistently outperforming carrier-only treated discs. Histological assessments supported these findings, showing trends toward extracellular matrix restoration in both treatment groups. While both biomaterials demonstrated reparative potential, HA showed greater consistency in supporting NP cells in promoting disc regeneration. These results underscore HA’s potential as a superior carrier for NP cell-based therapies in addressing disc degeneration. Full article

Low back pain (LBP) is a significant global health issue, contributing to disability and socioeconomic burdens worldwide. The degeneration of the human intervertebral disc (IVD) is a critical factor in the pathogenesis of LBP. Recent studies have emphasized the significance of a specific set of genes and extracellular matrix (ECM) in IVD health. In particular, Noggin has emerged as a critical gene due to its high expression levels in healthy nucleus pulposus cells (NPCs) observed in our previous research. In this study, it was hypothesized that decreased Noggin expression in NPCs is associated with IVD degeneration and contributes to LBP development. A lentivirus-mediated RNAi was applied to knock down Noggin expression in primary NPCs from six human donors. The NPCs after transduction were evaluated through cell viability analysis, XTT assay, and cell apoptosis analyses. After two weeks, a colony formation assay was used to examine the anchor-independent growth ability of transduced cells. At the transcript level, anabolic and catabolic markers were quantified using RT-qPCR. The results demonstrated that lentivirus-mediated downregulation of Noggin significantly inhibited cell proliferation, reduced cell viability, and suppressed colony formation, while inducing apoptosis in human NPCs in vitro. Notably, it disrupted cellular anabolic processes and promoted catabolic activity in human NPCs post-transduction. Our findings indicated that the degeneration of human IVD is possibly related to decreased Noggin expression in NPCs. This research provides valuable insights into the role of Noggin in IVD homeostasis and its implications in LBP treatment. Full article

Intervertebral disc degeneration (IDD) progresses owing to damage and depletion of nucleus pulposus (NP) cells. Cytoprotection mitigates oxidative stress, nutrient deprivation, and mechanical stress, which lead to cell damage and necrosis. We aimed to examine the protective effect of Raphanus sativus Linne (RSL), common radish, against oxidative stress by H₂O₂ in human NP cells and whether the RSL extracts can inhibit triggering receptor expressed on myeloid cells 2 (TREM2), an inducer of apoptosis and degeneration in NP cells. We administered hydrogen peroxide (H₂O₂) to cultured human NP cells treated with RSL extracts. We used immunoblotting and quantitative PCR to investigate expression of the apoptosis-associated proteins in cultured cells. RSL significantly enhanced cell survival by suppressing the activation of cleaved caspase-3 and Bax. In contrast, RSL extract increased Bcl2 concentration to downregulate apoptosis. Additionally, RSL treatment notably enhanced the mRNA levels of ACAN and Col2a1 while significantly reducing those of ADAMTS-4, ADAMTS-5, MMP3, and MMP13, key genes involved in NP degeneration. While H₂O₂ elevated TREM2 expression, causing disc degeneration, RSL downregulated TREM2 expression. Thus, our findings imply that RSL supports human NP cells under oxidative stress and regulates the pathways underlying disc degeneration, particularly TREM2, and that RSL extracts may potentially prevent IDD. Full article

Cell transplantation is being actively explored as a regenerative therapy for discogenic back pain. This study explored the regenerative potential of Tie2⁺ nucleus pulposus progenitor cells (NPPCs) from intervertebral disc (IVD) tissues derived from young (<25 years of age) and old (>60 years of age) patient donors. We employed an optimized culture method to maintain Tie2 expression in NP cells from both donor categories. Our study revealed similar Tie2 positivity rates regardless of donor types following cell culture. Nevertheless, clear differences were also found, such as the emergence of significantly higher (3.6-fold) GD2 positivity and reduced (2.7-fold) proliferation potential for older donors compared to young sources. Our results suggest that, despite obtaining a high fraction of Tie2⁺ NP cells, cells from older donors were already committed to a more mature phenotype. These disparities translated into functional differences, influencing colony formation, extracellular matrix production, and in vivo regenerative potential. This study underscores the importance of considering age-related factors in NPPC-based therapies for disc degeneration. Further investigation into the genetic and epigenetic alterations of Tie2⁺ NP cells from older donors is crucial for refining regenerative strategies. These findings shed light on Tie2⁺ NPPCs as a promising cell source for IVD regeneration while emphasizing the need for comprehensive understanding and scalability considerations in culture methods for broader clinical applicability. Full article