Progression Of Intervertebral Disc Degeneration Research Articles

Integrating Bulk RNA and Single-Cell RNA Sequencing Identifies and Validates Lactylation-Related Signatures for Intervertebral Disc Degeneration.

Glycolysis-related lactylation has gained wide attention for regulating various cellular functions and diseases. Nevertheless, its intricate involvement in intervertebral disc degeneration (IVDD) is not yet fully understood. In this study, we unrevealed the intricate association between elevated lactylation levels and the development of IVDD. Subsequently, we harvested the lactylation-related genes (LRGs) and systematically analysed the expression levels of these genes to establish a gene signature related to IVDD through multiple bulk RNA sequencing data. Six hub LRGs were determined and closely associated with the increased severity of IVDD. Among the six genes, CBX3 was the most upregulated in both invivo and invitro experiments. Furthermore, molecular docking identified atosiban acetate as a specific inhibitor for CBX3, and inhibiting the expression of CBX3 using atosiban acetate significantly repressed the glycolysis activity and global lactylation level, thus alleviating the progression of IVDD. In conclusion, the lactylation correlates positively with IVDD and the LRG signature could be used as a biomarker for the effective clinical treatment of IVDD. CBX3 emerged as one of the key LRGs in IVDD, and atosiban acetate, as a specific inhibitor for CBX3, may be a promising therapeutic candidate for IVDD by affecting lactylation.

Oxygen-controllable injectable hydrogel alleviates intervertebral disc degeneration by balancing extracellular matrix metabolism

Nucleus pulposus (NP) cells, situated at the core of intervertebral discs, have acclimated to a hypoxic environment, orchestrating the equilibrium of extracellular matrix metabolism (ECM) under the regulatory influence of hypoxia inducible factor-1α (HIF-1α). Neovascularization and increased oxygen content pose a threat, triggering ECM degradation and intervertebral disc degeneration (IVDD). To address this, our study devised an oxygen-controllable strategy, introducing laccase into an injectable and ultrasound-responsive gelatin/agarose hydrogel. Laccase-mediated reactions were employed to deplete oxygen, establishing a hypoxic microenvironment that upregulated HIF-1α expression. The activation of hypoxia-inducible factors significantly enhanced the expression of aggrecan and collagen II, concurrently suppressing Matrix metalloproteinases (MMP13) and A Disintegrin and Metalloproteinase with Thrombospondin motifs (ADAMTS5) levels, thereby restoring the equilibrium of ECM metabolism. Simultaneously, the hydrogel facilitated the recruitment of stem cells into the NP through the controlled release of ATI2341, activating C-X-C chemokine receptor type 4 (CXCR4). Moreover, ultrasound amplification enhanced ATI2341 release, promoting the migration of NP stem cells. The hydrogel's efficacy in mitigating metabolic imbalances and inhibiting IVDD progression was substantiated in a rat puncture IVDD model through hydrogel injection into the discs. In conclusion, this hypoxia-inducible hydrogel, responsive to thermal stimuli from ultrasound, presents a promising avenue for IVDD treatment.

PLGA/BK microspheres targeting the bradykinin signaling pathway as a therapeutic strategy to delay intervertebral disc degeneration

Intervertebral disc degeneration(IVDD) is a common spinal condition with limited effective treatments available. This study aims to investigate the impact of poly(lactic-co-glycolic acid)/Bradykinin (PLGA/BK) microspheres on IVDD and its underlying mechanisms. We collected nucleus pulposus samples from both healthy and degenerated human intervertebral disks and conducted immunohistochemical analyses, revealing reduced BK expression in degenerated tissues. Subsequently, we used BK to treat nucleus pulposus cells and conducted Bulk RNA sequencing (RNA-seq), identifying BK’s involvement in cellular senescence, extracellular matrix metabolism, and the PI3K signaling pathway. Further experiments using tert-butyl hydroperoxide (TBHP)-induced cell senescence showed that BK treatment reduced senescence, enhanced extracellular matrix synthesis, and inhibited degradation, along with activation of the PI3K pathway. These effects were mediated through B2R (BK receptor 2) and the downstream PI3K pathway. Following this, we developed sustained-release BK microspheres with an optimized manufacturing process. In vitro co-culture experiments showed no observable toxicity. We established an IVDD model in rat tail vertebrae through fine needle puncture, administering local injections of BK sustained-release microspheres. Using various experimental methods, including X-ray, MRI, histopathology, and immunohistochemistry, we found that these microspheres could slow the progression of IVDD. This study highlights the potential of injectable PLGA/BK microspheres to regulate cellular senescence and extracellular matrix metabolism via the B2R and PI3K pathways, ultimately delaying IVDD.

Melatonin attenuates degenerative disc degression by downregulating DLX5 via the TGF/Smad2/3 pathway in nucleus pulposus cells.

Intervertebral disc degeneration (IVDD) is the leading cause of low back pain, and apoptosis plays a key role in its pathogenesis. Distal-less homeobox 5 (Dlx5) has been reported to induce cell apoptosis. Melatonin, as a powerful antiapoptotic agent, has been widely reported. This study aimed to investigate the role of DLX5 in the pathogenesis of IVDD and the potential therapeutic role of melatonin in targeting DLX5 in IVDD. Western blotting, RT-qPCR, immunohistochemistry, si-DLX5, Ex-DLX5, flow cytometry, and immunofluorescence were used to examine the regulatory effect of DLX5 on apoptosis. Therapeutic efficacy was assessed by the intraperitoneal injection of melatonin into IVDD mice. The expression level of DLX5 is significantly increased in IVDD, and the expression levels were positively correlated with the grade of IVDD. DLX5 was significantly upregulated in TNF-α-induced degenerative NP cells. Degenerative NP cells transfected with si-DLX5 exhibited significantly less apoptosis than control cells. Melatonin significantly alleviated IVDD in surgically induced IVDD model mice. The results revealed that the expression of DLX5 was positively correlated with the severity of IVDD and that melatonin ameliorated DLX5-induced apoptosis and extracellular matrix imbalance by inhibiting the TGF-β/Smad signaling pathway. This study may provide therapeutic strategies to alleviate inflammation-induced apoptosis IVDD-associated inflammation-induced apoptosis. DLX5 plays an important role in IVDD progression by promoting apoptosis, and melatonin represents a promising therapeutic strategy for alleviating IVDD-associated inflammation and apoptosis.

Single-cell sequencing reveals cellular heterogeneity of nucleus pulposus in intervertebral disc degeneration

The nucleus pulposus (NP) plays a vital role in intervertebral disc degeneration (IVDD). Previous studies have revealed cellular heterogeneity in NP tissue during IVDD progression. However, the cellular and molecular alterations of diverse cell clusters during IVDD remain to be fully elucidated. NP tissues were isolated from patients with different grades of IVDD undergoing discectomy, and then subjected to single-cell RNA sequencing (scRNA-seq). Cell subsets were identified based on unbiased clustering of gene expression profiles. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed to determine the molecular features of diverse cell clusters. Monocle analysis was used to illustrate the differentiation trajectories of chondrocytes. Additionally, CellPhoneDB analysis revealed potential interactions between chondrocytes and other cells during IVDD. Based on the expression profiles of 47,610 individual cells, eight putative clusters including chondrocytes, endothelial cells, fibroblasts, macrophages, mural cells, osteoclasts, proliferating stromal cells and T cells were identified. The chondrocyte cluster was classified into three subsets, C1-C3, which were associated with stress-resistance, fibrosis and inflammatory responses, respectively. Pseudo-time trajectories suggested that chondrocytes gradually differentiated into fibroblasts during IVDD. Immune cells including cDC2s, macrophages and monocytes were identified. Further analysis showed that chondrocytes might communicate with immune cells via the MIF, TNFSF9, SPP1 and CCL4L2 signaling pathways. In addition, we found that invading endothelial cells might interact with chondrocytes through the COL4A1, CXCL12, VEGFA and SEMA3E signaling pathways. Our results reveal the cellular complexity and phenotypic characteristics of NP tissues at single-cell resolution, which will contribute to the in-depth investigation of preventative and regenerative strategies for IVDD.

Identification of extracellular matrix proteins in plasma as a potential biomarker for intervertebral disc degeneration.

Recently, there has been significant focus on extracellular matrix proteolysis due to its importance in the pathological progression of intervertebral disc degeneration (IVDD). The present study investigates the circulating levels of extracellular matrix proteins in the plasma of IVDD and determines their potential relevance as biomarkers in disc degeneration. Global proteomic analysis was performed in the plasma samples of 10 healthy volunteers (HV) and 10 diseased subjects (DS) after depletion of highly abundant proteins such as albumin and IgG. We identified 144 and 135 matrix-associated proteins in plasma samples from healthy volunteers (HV) and patients with disc degeneration (DS), respectively. Among these, 49 of the matrix-associated proteins were identical to the proteins found in intervertebral disc (IVD) tissues retrieved from the in-house library. Applying stringent parameters, we selected 28 proteins, with 26 present in DS and 21 in HV. 19 proteins were found common between the groups, two of which-aggrecan (ACAN) and fibulin 1 (FBLN1) - showed statistically significant differences. Specifically, ACAN was up-regulated and FBLN1 was down-regulated in the DS-plasma. In particular, DS-plasma exhibited specific expression of collagen type 2a1 (COL2A1), native to the nucleus pulposus. The distinct presence of collagen type 2a1 and the elevated expression of aggrecan in IVDD plasma may serve as the basis for the development of a potential biomarker for monitoring the progression of disc degeneration.

Unveiling the Gut-Disc Axis: How Microbiome Dysbiosis Accelerates Intervertebral Disc Degeneration.

The gut microbiome (GM), often referred to as the second genome of the human body, plays a crucial role in various metabolic processes and mediates the development of numerous diseases. Intervertebral disc degeneration (IDD) is an age-related degenerative spinal disease characterized by the loss of disc height, hydration, and integrity, leading to pain and reduced mobility. Although the pathogenesis of IDD is not fully understood, recent studies suggest that dysbiosis of the gut microbiome may accelerate the progression of IDD through multiple mechanisms. This article begins by discussing the potential relationship between GM dysbiosis and human diseases, followed by a comprehensive review of the regulatory mechanisms of GM in skeletal diseases within the gut-disc axis framework. Furthermore, it explores three potential pathways through which GM dysbiosis may mediate the development of IDD: immunomodulation, bacterial translocation and colonization, and the decomposition and absorption of intestinal metabolites. These pathways can disrupt disc cell homeostasis and promote degenerative changes. Finally, this paper summarizes for the first time the potential therapeutic approaches for delaying IDD by targeting the gut-disc axis, providing new insights into the pathogenesis and regenerative repair strategies for IDD.

The Role of Microvascular Variations in the Process of Intervertebral Disk Degeneration and Its Regulatory Mechanisms: A Literature Review.

Microvascular changes are considered key factors in the process of intervertebral disk degeneration (IDD). Microvascular invasion and growth into the nucleus pulposus (NP) and cartilaginous endplates are unfavorable factors that trigger IDD. In contrast, the rich distribution of microvessels in the bony endplates and outer layers of the annulus fibrosus is an important safeguard for the nutrient supply and metabolism of the intervertebral disk (IVD). In particular, the adequate supply of microvessels in the bony endplates is the main source of the nutritional supply for the entire IVD. Microvessels can affect the progression of IDD through a variety of pathways. Many studies have explored the effects of microvessel alterations in the NP, annulus fibrosus, cartilaginous endplates, and bony endplates on the local microenvironment through inflammation, apoptosis, and senescence. Studies also elucidated the important roles of microvessel alterations in the process of IDD, as well as conducted in-depth explorations of cytokines and biologics that can inhibit or promote the ingrowth of microvessels. Therefore, the present manuscript reviews the published literature on the effects of microvascular changes on IVD to summarize the roles of microvessels in IVD and elaborate on the mechanisms of action that promote or inhibit de novo microvessel formation in IVD.

From structure to therapy: the critical influence of cartilaginous endplates and microvascular network on intervertebral disc degeneration.

The intervertebral disc (IVD) is the largest avascular structure in the human body. The cartilaginous endplate (CEP) is a layer of translucent cartilage located at the upper and lower edges of the vertebral bodies. On one hand, CEPs endure pressure from within the IVD and the tensile and shear forces of the annulus fibrosus, promoting uniform distribution of compressive loads on the vertebral bodies. On the other hand, microvascular diffusion channels within the CEP serve as the primary routes for nutrient supply to the IVD and the transport of metabolic waste. Degenerated CEP, characterized by increased stiffness, decreased permeability, and reduced water content, impairs substance transport and mechanical response within the IVD, ultimately leading to intervertebral disc degeneration (IDD). Insufficient nutrition of the IVD has long been considered the initiating factor of IDD, with CEP degeneration regarded as an early contributing factor. Additionally, CEP degeneration is frequently accompanied by Modic changes, which are common manifestations in the progression of IDD. Therefore, this paper comprehensively reviews the structure and physiological functions of CEP and its role in the cascade of IDD, exploring the intrinsic relationship between CEP degeneration and Modic changes from various perspectives. Furthermore, we summarize recent potential therapeutic approaches targeting CEP to delay IDD, offering new insights into the pathological mechanisms and regenerative repair strategies for IDD.

PRDM1 promotes nucleus pulposus cell pyroptosis leading to intervertebral disc degeneration via activating CASP1 transcription.

Intervertebral disc degeneration (IVDD) is a primary contributor to low back pain and poses a considerable burden to society. However, the molecular mechanisms underlying IVDD remain to be elucidated. PR/SET domain 1 (PRDM1) regulates cell proliferation, apoptosis, and inflammatory responses in various diseases. Despite these regulatory functions, the mechanism of action of PRDM1 in IVDD remains unexplored. In this study, we investigated the role and underlying mechanisms of action of PRDM1 in IVDD progression. The expression of PRDM1 in nucleus pulposus (NP) tissues and NP cells (NPCs) was assessed using western blotting, immunohistochemistry, and immunofluorescence. The effects of PRDM1 on IVDD progression were investigated in vitro and in vivo. Mechanistically, mRNA sequencing, chromatin immunoprecipitation, and dual-luciferase reporter assays were performed to confirm that PRDM1 triggered CASP1 transcription. Our study demonstrated for the first time that PRDM1 expression was substantially upregulated in degenerated NP tissues and NPCs. PRDM1 overexpression promoted NPCs pyroptosis by inhibiting mitophagy and exacerbating IVDD progression, whereas PRDM1 silencing exerted the opposite effect. Furthermore, PRDM1 activated CASP1 transcription, thereby promoting NPCs pyroptosis in vitro. Notably, CASP1 silencing reversed the effects of PRDM1 on the NPCs. To the best of our knowledge, this study is the first to demonstrate that PRDM1 silencing inhibits NPCs pyroptosis by repressing CASP1 transcription, which may be a promising new therapeutic target for IVDD.

Selenium nanoparticles ameliorate lumbar disc degeneration by restoring GPX1-mediated redox homeostasis and mitochondrial function of nucleus pulposus cells

Intervertebral disc degeneration (IVDD) is a prevalent musculoskeletal disorder that involves the excessive accumulation of reactive oxygen species (ROS), resulting in mitochondrial dysfunction and matrix metabolism imbalance in nucleus pulposus cells (NPCs). Selenium, an indispensable trace element, plays a crucial role in maintaining mitochondrial redox homeostasis by being incorporated into antioxidant selenoproteins as selenocysteine. In this study, we employed a straightforward synthesis method to produce selenium nanoparticles (SeNPs) with consistent size and distribution, and evaluated their potential protective effects in ameliorating IVDD. In a simulated inflammatory environment induced by interleukin-1beta (IL-1β) in vitro, SeNPs demonstrated a protective effect on the matrix synthesis capacity of NPCs through the up-regulation of aggrecan and type II collagen, while concurrently suppressing the expression of matrix degradation enzymes including MMP13 and ADAMTS5. Additionally, SeNPs preserved mitochondrial integrity and restored impaired mitochondrial energy metabolism by activating glutathione peroxidase1 (GPX1) to rebalance redox homeostasis. In a rat lumbar disc model induced by puncture, the local administration of SeNPs preserved the hydration of nucleus pulposus tissue, promoted matrix deposition, and effectively mitigated the progression of IVDD. Our results indicate that the enhancement of GPX1 by SeNPs may offer a promising therapeutic approach for IVDD by restoring mitochondrial function and redox homeostasis.

Inactivation of Tnf-α/Tnfr signaling attenuates progression of intervertebral disc degeneration in mice.

Intervertebral disc degeneration (IVDD) is a major cause of low back pain (LBP), worsened by chronic inflammatory processes associated with aging. Tumor necrosis factor alpha (Tnf-α) and its receptors, Tnf receptor type 1 (Tnfr1) and Tnf receptor type 2 (Tnfr2), are upregulated in IVDD. However, its pathologic mechanisms remain poorly defined. To investigate the role of Tnfr in IVDD, we generated global Tnfr1/2 double knockout (KO) mice and age-matched control C57BL/6 male mice, and analyzed intervertebral disc (IVD)-related phenotypes of both genotypes under physiological conditions, aging, and lumbar spine instability (LSI) model through histological and immunofluorescence analyses and μCT imaging. Expression levels of key extracellular matrix (ECM) proteins in aged and LSI mice, especially markers of cell proliferation and apoptosis, were evaluated in aged (21-month-old) mice. At 4 months, KO and control mice showed no marked differences of IVDD-related parameters. However, at 21 months of age, the loss of Tnfr expression significantly alleviated IVDD-like phenotypes, including a significant increase in height of the nucleus pulposus (NPs) and reductions of endplates (EPs) porosity and histopathological scores, when compared to controls. Tnfr deficiency promoted anabolic metabolism of the ECM proteins and suppressed ECM catabolism. Tnfr loss largely inhibited hypertrophic differentiation, and, in the meantime, suppressed cell apoptosis and cellular senescence in the annulus fibrosis, NP, and EP tissues without affecting cell proliferation. Similar results were observed in the LSI model, where Tnfr deficiency significantly alleviated IVDD and enhanced ECM anabolic metabolism while suppressing catabolism. The deletion of Tnfr mitigates age-related and LSI-induced IVDD, as evidenced by preserved IVD structure, and improved ECM integrity. These findings suggest a crucial role of Tnf-α/Tnfr signaling in IVDD pathogenesis in mice. Targeting this pathway may be a novel strategy for IVDD prevention and treatment.

Isoliquiritigenin mitigates intervertebral disc degeneration induced by oxidative stress and mitochondrial impairment through a PPARγ-dependent pathway

ObjectivesOxidative stress, mitochondrial dysfunction, and apoptosis play significant roles in the degradation of extracellular matrix (ECM) in nucleus pulposus cells (NPCs), ultimately contributing to intervertebral disc degeneration (IVDD). This study investigates the potential of isoliquiritigenin (ISL), a natural extract known for its antioxidant, anti-inflammatory, and anti-atherosclerotic properties, to alleviate IVDD. MethodsThe viability of NPCs treated with ISL and tert-butyl hydroperoxide (TBHP) was assessed using the CCK-8 assay. Various techniques, including Western blot, qRT-PCR, immunofluorescence (IF), and immunohistochemistry, were employed to measure the expression of ECM components, oxidative stress markers, and apoptosis-related proteins. Mitochondrial function was evaluated through Western blot and IF analyses. Network pharmacology predicted ISL targets, and the expression levels of PPARγ were assessed using the aforementioned methods. The role of PPARγ in the therapeutic effects of ISL on IVDD was examined through siRNA knockdown. The therapeutic impact of ISL on puncture-induced IVDD in rats was evaluated using X-ray, MRI, and histological staining techniques. ResultsIn vitro, ISL reduced oxidative stress in NPCs, restored mitochondrial function, inhibited apoptosis, and improved the ECM phenotype. In vivo, ISL slowed the progression of IVDD in a rat model. Further analysis revealed that ISL enhances PPARγ activity and promotes its expression by direct binding, contributing to the delay of IVDD progression. ConclusionThis study demonstrates that ISL effectively treats puncture-induced IVDD in rats by inhibiting oxidative stress, restoring mitochondrial function, and reducing NPC apoptosis through a PPARγ-dependent mechanism. By balancing ECM synthesis and degradation, ISL presents a novel therapeutic approach for IVDD and identifies a promising target for treatment.

Hydrogel and Microgel Collaboration for Spatiotemporal Delivery of Biofactors to Awaken Nucleus Pulposus-Derived Stem Cells for Endogenous Repair of Disc.

Depletion of nucleus pulposus-derived stem cells (NPSCs) is a major contributing factor to the attenuation of endogenous regenerative capacity in intervertebral disc degeneration (IVDD). Introducing a hydrogel drug delivery system is a potential strategy for counteracting endogenous cell depletion. The present study proposes a delivery platform for the spatiotemporal release of multiple drugs by combining sodium alginate hydrogels with gelatin microgels (SCGP hydrogels). The SCGP hydrogels facilitated the initial release of chondroitin sulfate (ChS) and the gradual release of an independently developed parathyroid hormone-related peptide (P2). The combined action of these two small molecule drugs "awakened" the reserve NPSCs, mitigated cell damage induced by H2O2, significantly enhanced their biological activity, and promoted their differentiation toward nucleus pulposus cells. The mechanical and viscoelastic properties of the hydrogel are enhanced by physical and chemical dual cross-linking to adapt to the loading environment of the degenerated disc. A rat IVDD model is used to validate that the SCGP hydrogel can significantly inhibit the progression of IVDD and stimulate the endogenous repair of IVDD. Therefore, the spatiotemporal differential drug delivery system of the SCGP hydrogel holds promise as a convenient and efficacious therapeutic strategy for minimally invasive IVDD treatment.

The JNK signaling pathway in intervertebral disc degeneration.

Intervertebral disc degeneration (IDD) serves as the underlying pathology for various spinal degenerative conditions and is a primary contributor to low back pain (LBP). Recent studies have revealed a strong correlation between IDD and biological processes such as Programmed Cell Death (PCD), cellular senescence, inflammation, cell proliferation, extracellular matrix (ECM) degradation, and oxidative stress (OS). Of particular interest is the emerging evidence highlighting the significant involvement of the JNK signaling pathway in these fundamental biological processes of IDD. This paper explores the potential mechanisms through the JNK signaling pathway influences IDD in diverse ways. The objective of this article is to offer a fresh perspective and methodology for in-depth investigation into the pathogenesis of IDD by thoroughly examining the interplay between the JNK signaling pathway and IDD. Moreover, this paper summarizes the drugs and natural compounds that alleviate the progression of IDD by regulating the JNK signaling pathway. This paper aims to identify potential therapeutic targets and strategies for IDD treatment, providing valuable insights for clinical application.

Identification and validation of ferroptosis-related biomarkers in intervertebral disc degeneration.

Ferroptosis plays a significant role in intervertebral disc degeneration (IDD). Understanding the key genes regulating ferroptosis in IDD could reveal fundamental mechanisms of the disease, potentially leading to new diagnostic and therapeutic targets. Public datasets (GSE23130 and GSE70362) and the FerrDb database were analyzed to identify ferroptosis-related genes (DE-FRGs) involved in IDD. Single-cell RNA sequencing data (GSE199866) was used to validate the specific roles and expression patterns of these genes. Immunohistochemistry and Western blot analyses were subsequently conducted in both clinical samples and mouse models to assess protein expression levels across different tissues. The analysis identified seven DE-FRGs, including MT1G, CA9, AKR1C1, AKR1C2, DUSP1, CIRBP, and KLHL24, with their expression patterns confirmed by single-cell RNA sequencing. Immunohistochemistry and Western blot analysis further revealed that MT1G, CA9, AKR1C1, AKR1C2, DUSP1, and KLHL24 exhibited differential expression during the progression of IDD. Additionally, the study highlighted the potential immune-modulatory functions of these genes within the IDD microenvironment. Our study elucidates the critical role of ferroptosis in IDD and identifies specific genes, such as MT1G and CA9, as potential targets for diagnosis and therapy. These findings offer new insights into the molecular mechanisms underlying IDD and present promising avenues for future research and clinical applications.

A novel spherical GelMA-HAMA hydrogel encapsulating APET×2 polypeptide and CFIm25-targeting sgRNA for immune microenvironment modulation and nucleus pulposus regeneration in intervertebral discs

MethodsSingle-cell transcriptomics and high-throughput transcriptomics were used to screen factors significantly correlated with intervertebral disc degeneration (IDD). Expression changes of CFIm25 were determined via RT-qPCR and Western blot. NP cells were isolated from mouse intervertebral discs and induced to degrade with TNF-α and IL-1β. CFIm25 was knocked out using CRISPR-Cas9, and CFIm25 knockout and overexpressing nucleus pulposus (NP) cell lines were generated through lentiviral transfection. Proteoglycan expression, protein expression, inflammatory factor expression, cell viability, proliferation, migration, gene expression, and protein expression were analyzed using various assays (alcian blue staining, immunofluorescence, ELISA, CCK-8, EDU labeling, transwell migration, scratch assay, RT-qPCR, Western blot). The GelMA-HAMA hydrogel loaded with APET×2 polypeptide and sgRNA was designed, and its effects on NP regeneration were assessed through in vitro and mouse model experiments. The progression of IDD in mice was evaluated using X-ray, H&E staining, and Safranin O-Fast Green staining. Immunohistochemistry was performed to determine protein expression in NP tissue. Proteomic analysis combined with in vitro and in vivo experiments was conducted to elucidate the mechanisms of hydrogel action.ResultsCFIm25 was upregulated in IDD NP tissue and significantly correlated with disease progression. Inhibition of CFIm25 improved NP cell degeneration, enhanced cell proliferation, and migration. The hydrogel effectively knocked down CFIm25 expression, improved NP cell degeneration, promoted cell proliferation and migration, and mitigated IDD progression in a mouse model. The hydrogel inhibited inflammatory factor expression (IL-6, iNOS, IL-1β, TNF-α) by targeting the p38/NF-κB signaling pathway, increased collagen COLII and proteoglycan Aggrecan expression, and suppressed NP degeneration-related factors (COX-2, MMP-3).ConclusionThe study highlighted the crucial role of CFIm25 in IDD and introduced a promising therapeutic strategy using a porous spherical GelMA-HAMA hydrogel loaded with APET×2 polypeptide and sgRNA. This innovative approach offers new possibilities for treating degenerated intervertebral discs.Graphical Molecular mechanism of GCA targeting CFIm25 in NP cells to promote regeneration of degenerated intervertebral disc NP

Melatonin mitigates matrix stiffness-induced intervertebral disk degeneration by inhibiting reactive oxygen species and melatonin receptors mediated PI3K/AKT/NF-κB pathway.

Intervertebral disk degeneration (IVDD) may lead to an increase in extracellular matrix (ECM) stiffness, potentially contributing to the progression of the disease. Melatonin reportedly mitigates IVDD; however, its potential to attenuate elevated matrix stiffness-induced IVDD remains unexplored. Therefore, we aimed to investigate whether melatonin can alleviate the progression of IVDD triggered by increased matrix stiffness and elucidate its underlying mechanisms. Nucleus pulposus (NP) tissues were collected from patients, and ECM stiffness, reactive oxygen species (ROS) levels, apoptosis rates, and P65 expression in these tissues with varying Pfirrmann scores were determined. In vitro experiments were conducted to investigate the effects of melatonin on various pathophysiological mechanisms within the NP cells cultured on soft substrates with differing stiffness levels. Our findings revealed a positive correlation between ECM stiffness in human NP tissue and degree of IVDD. In addition, phosphorylation of P65 exhibited a strong association with matrix stiffness. Enhanced levels of ROS and cellular apoptosis were observed within degenerated intervertebral disks. In vitro experiments demonstrated that melatonin significantly inhibited catabolism and apoptosis induced by stiff matrices, along with elevated ROS levels. Furthermore, we observed that melatonin inhibited NP cell catabolism and apoptosis by reducing the melatonin receptors mediated activation of the PI3K/AKT and nuclear factor-kappa B (NF-κB) pathways. Also, we found that the reduction of ROS by melatonin can assist in inhibiting the activation of the NF-κB pathway. The outcomes of the in vivo experiments corroborated the results of the in vitro experiments, illustrating that melatonin treatment could alleviate the compression-induced upregulation of matrix stiffness in NP and IVDD. Collectively, melatonin can potentially alleviate high matrix stiffness-induced IVDD by reducing intracellular ROS levels and inhibiting the PI3K/AKT/NF-κB pathway.NEW & NOTEWORTHY Melatonin mitigates intervertebral disk degeneration (IVDD) induced by matrix stiffness through reactive oxygen species (ROS) reduction. Matrix stiffness is related to increased nucleus pulposus cell ROS, apoptosis, and degeneration. Melatonin inhibits PI3K/AKT/NF-κB pathways via melatonin receptors in a stiff matrix environment. In vivo, melatonin restores disk height and alleviates IVDD progression.

Causal relationship between systemic circulatory inflammatory regulators and intervertebral disc degeneration: A bidirectional 2-sample Mendelian randomization study.

In the context of the development of intervertebral disc degeneration (IDD), inflammatory mediators play a pivotal role. Nevertheless, due to the influence of the inflammatory microenvironment, the causal relationship between specific inflammatory mediators and the development of IDD remains uncertain. The understanding of the causal relationship between inflammatory mediators and IDD is of great importance in preventing and delaying disc degeneration in the future. We utilized genetic data concerning systemic circulating inflammatory regulators obtained from a Genome-Wide Association Study (GWAS) analyzing 41 serum cytokines in a cohort of 8293 individuals from Finland. The genetic data for IDD were derived from the most recent GWAS summary statistics conducted within the FinnGen consortium, encompassing 37,636 IDD cases and 270,964 controls. Our analysis employed bidirectional 2-sample Mendelian randomization (MR) techniques, which included several MR methods such as MR Egger, weighted median, inverse variance weighted, weighted mode, and simple mode. Additionally, the MR-PRESSO method was employed to identify horizontal pleiotropy, heterogeneity was quantified using the Cochran Q statistic, and MR-Egger intercept analysis was performed to assess pleiotropy. We established causal relationships between 3 specific inflammatory factors and IDD. Elevated levels of MIP-1β (OR = 0.956, 95% CI: -0.08 to -0.006; P = .02) and IFN-G (OR = 0.915, 95% CI: -0.16 to -0.02; P = .01) expression were associated with a reduced risk of IDD. Conversely, genetic susceptibility to IDD was linked to a decrease in IL-13 levels (OR = 0.967, 95% CI: -0.063 to -0.004; P = .03). In this study, we have identified inflammatory factors that exhibit a causal relationship with the onset and progression of IDD, as supported by genetic predictions.

The treatment strategies for the repair of intervertebral disc degeneration

The traditional treatment of intervertebral disc degeneration (IVDD) mainly focuses on symptomatic treatment, and cannot restore the physiological structure and function of the intervertebral disc. Therefore, more and more scholars begin to pay attention to the application of regenerative medicine and its derived therapeutic methods in IVDD. From the histological perspective, the early stage of IVDD shows the imbalance between synthesis and catabolism, but the cell number and tissue structure are relatively complete, and the intervention of exogenous molecules or gene therapy can achieve extracellular matrix (ECM) regeneration. With the progress of IVDD, the replenishment of healthy cells is the key to treatment. In the final stage, the cell number and tissue structure are disordered. Biological materials with certain mechanical strength and cell load can be used to supplement ECM and healthy cells to realize the repair and regeneration of IVDD. Molecular, cell and gene therapy, combined with the application of new biomaterials, the treatment of IVDD is more inclined to compensate for the shortcomings through a combination approach in the future, in order to achieve the purpose of repair and regeneration.