Palmitic acid induces lipid droplet accumulation and senescence in nucleus pulposus cells via ER-stress pathway.

Intervertebral disc degeneration (IDD) is a highly prevalent musculoskeletal disorder affecting millions of adults worldwide, but a poor understanding of its pathogenesis has limited the effectiveness of therapy. In the current study, we integrated untargeted LC/MS metabolomics and magnetic resonance spectroscopy data to investigate metabolic profile alterations during IDD. Combined with validation via a large-cohort analysis, we found excessive lipid droplet accumulation in the nucleus pulposus cells of advanced-stage IDD samples. We also found abnormal palmitic acid (PA) accumulation in IDD nucleus pulposus cells, and PA exposure resulted in lipid droplet accumulation and cell senescence in an endoplasmic reticulum stress-dependent manner. Complementary transcriptome and proteome profiles enabled us to identify solute carrier transporter (SLC) 43A3 involvement in the regulation of the intracellular PA level. SLC43A3 was expressed at low levels and negatively correlated with intracellular lipid content in IDD nucleus pulposus cells. Overexpression of SLC43A3 significantly alleviated PA-induced endoplasmic reticulum stress, lipid droplet accumulation and cell senescence by inhibiting PA uptake. This work provides novel integration analysis-based insight into the metabolic profile alterations in IDD and further reveals new therapeutic targets for IDD treatment.

Genkwanin alleviates intervertebral disc degeneration via regulating ITGA2/PI3K/AKT pathway and inhibiting apoptosis and senescence.

Intervertebral disc degeneration (IVDD) is a progressive degenerative disease influenced by various factors. Genkwanin, a known anti-inflammatory flavonoid, has not been explored for its potential in IVDD management. This study aims to investigate the effects and mechanisms of genkwanin on IVDD. In vitro, cell experiments revealed that genkwanin dose-dependently inhibited Interleukin-1ß-induced expression levels of inflammatory factors (Interleukin-6, inducible nitric oxide synthase, cyclooxygenase-2) and degradation metabolic protein (matrix metalloproteinase-13). Concurrently, genkwanin upregulated the expression of synthetic metabolism genes (type II collagen, aggrecan). Moreover, genkwanin effectively reduced the phosphorylation of phosphatidylinositol 3-kinase (PI3K)/AKT/mammalian target of rapamycin, mitogen-activated protein kinase (MAPK), and nuclear factor-κB (NF-κB) pathways. Transcriptome sequencing analysis identified integrin α2 (ITGA2) as a potential target of genkwanin, and silencing ITGA2 reversed the activation of PI3K/AKT pathway induced by Interleukin-1ß. Furthermore, genkwanin alleviated Interleukin-1ß-induced senescence and apoptosis in nucleus pulposus cells. In vivo animal experiments demonstrated that genkwanin mitigated the progression of IVDD in the rat model through imaging and histological examinations. In conclusion, This study suggest that genkwanin inhibits inflammation in nucleus pulposus cells, promotes extracellular matrix remodeling, suppresses cellular senescence and apoptosis, through the ITGA2/PI3K/AKT, NF-κB and MAPK signaling pathways. These findings indicate that genkwanin may be a promising therapeutic candidate for IVDD.

Dynamics of CD44+ bovine nucleus pulposus cells with inflammation.

Intervertebral Disc (IVD) degeneration has been associated with a chronic inflammatory response, but knowledge on the contribution of distinct IVD cells, namely CD44, to the progression of IVD degeneration remains elusive. Here, bovine nucleus pulposus (NP) CD44 cells were sorted and compared by gene expression and proteomics with the negative counterpart. NP cells were then stimulated with IL-1b (10 ng/ml) and dynamics of CD44 gene and protein expression was analyzed upon pro-inflammatory treatment. The results emphasize that CD44 has a multidimensional functional role in IVD metabolism, ECM synthesis and production of neuropermissive factors. CD44 widespread expression in NP was partially associated with CD14 and CD45, resulting in the identification of distinct cell subsets. In conclusion, this study points out CD44 and CD44-based cell subsets as relevant targets in the modulation of the IVD pro-inflammatory/degenerative cascade.

Effects of miR-129-5p on inflammation and nucleus pulposus cell apoptosis in rats with intervertebral disc degeneration through JNK signaling pathway.

This study aimed to explore the effects of miR-129-5p on inflammation and nucleus pulposus (NP) cell apoptosis in rats with intervertebral disc degeneration (IVDD) through the c-Jun N-terminal kinase (JNK) signaling pathway. A total of 20 rats were randomly divided into control group (n=10) or IVDD group (n=10). The mRNA expressions of miR-129-5p and apoptosis index Fas in IVDD tissues were determined using RT-PCR. NP cell apoptosis rate was detected via TUNEL assay. NP cells were extracted from IVDD tissues for primary culture. Subsequently, the cells were transfected with miR-129-5p inhibitor or mimic to inhibit or overexpress miR-129-5p, respectively. Furthermore, the changes in the JNK pathway indexes and apoptosis indexes were detected using Western blotting. In IVDD group, the expression of miR-129-5p was significantly down-regulated, while the transcriptional level of Fas was up-regulated compared with those in control group. Pearson correlation analysis revealed a negative correlation between the expressions of miR-129-5p and Fas mRNA (r=-0.75, P<0.05). IVDD group exhibited significantly higher levels of serum TNF-α, IL-6 and IL-1 than control group. Subsequent TUNEL assay indicated that the apoptosis rate was evidently higher in IVDD group (60.6%) than control group (2.5%). The results of Western blotting showed that the protein expressions of JNK1, JNK2 and Fas remarkably rose in IVDD group compared with those in control group. However, they declined remarkably in miR-129-5p mimic group compared with those in control group. Furthermore, such trends were significantly reversed in miR-129-5p inhibitor group. MiR-129-5p was significantly down-regulated in IVDD, whose overexpression has anti-inflammatory and anti-apoptotic effects.

Regulation of endoplasmic reticulum stress on autophagy and apoptosis of nucleus pulposus cells in intervertebral disc degeneration and its related mechanisms.

Intervertebral disc degeneration (IVDD) is a common and frequent disease in orthopedics, which seriously affects the quality of life of patients. Endoplasmic reticulum stress (ERS)-regulated autophagy and apoptosis play an important role in nucleus pulposus (NP) cells in IVDD. Hypoxia and serum deprivation were used to induce NP cells. Cell counting kit-8 (CCK-8) assay was used to detect cell activity and immunofluorescence (IF) was applied for the appraisement of glucose regulated protein 78 (GRP78) and green fluorescent protein (GFP)-light chain 3 (LC3). Cell apoptosis was detected by flow cytometry and the expression of LC3II/I was detected by western blot. NP cells under hypoxia and serum deprivation were induced by lipopolysaccharide (LPS), and intervened by ERS inhibitor (4-phenylbutyric acid, 4-PBA) and activator (Thapsigargin, TP). Then, above functional experiments were conducted again and western blot was employed for the evaluation of autophagy-, apoptosis and ERS-related proteins. Finally, NP cells under hypoxia and serum deprivation were stimulated by LPS and intervened using apoptosis inhibitor z-Val-Ala-DL-Asp-fluoromethyl ketone (Z-VAD-FMK) and autophagy inhibitor 3-methyladenine (3-MA). CCK-8 assay, IF, flow cytometry and western blot were performed again. Besides, the levels of inflammatory cytokines were measured with enzyme-linked immunosorbent assay (ELISA) and the protein expressions of programmed death markers were estimated with western blot. It showed that serum deprivation induces autophagy and apoptosis. ERS was significantly activated by LPS in hypoxic and serum deprivation environment, and autophagy and apoptosis were significantly promoted. Overall, ERS affects the occurrence and development of IVDD by regulating autophagy, apoptosis and other programmed death.

Single-cell RNA-seq analysis reveals the Wnt/Ca2+ signaling pathway with inflammation, apoptosis in nucleus pulposus degeneration.

BACKGROUND: Increasing studies have shown degeneration of nucleus pulposus cells (NPCs) as an critical part of the progression of intervertebral disc degeneration (IVDD). However, there are relatively few studies on single-cell transcriptome contrasts in human degenerated NPCs. Moreover, differences in Wnt/Ca2+ signaling in human degenerated nucleus pulposus cells have not been elucidated. The aim of this study is to investigate the differential expression of Wnt/Ca2+ signaling pathway between normal and degenerated nucleus pulposus cells in humans and try to investigate its mechanism. METHODS: We performed bioinformatics analysis using our previously published findings to construct single cell expression profiles of normal and degenerated nucleus pulposus. Then, in-depth differential analysis was used to characterize the expression of Wnt/Ca2+ signaling pathway between normal and degenerated nucleus pulposus cells in humans. RESULTS: The obtained cell data were clustered into five different chondrocytes clusters, which chondrocyte 4 and chondrocyte 5 mainly accounted for a high proportion in degenerated nucleus pulposus tissues, but rarely in normal nucleus pulposus tissues. Genes associated within the Wnt/Ca2+ signaling pathway, such as Wnt5B, FZD1, PLC (PLCB1), CaN (PPP3CA) and NAFATC1 are mainly present in chondrocyte 3, chondrocyte 4 and chondrocyte 5 from degenerated nucleus pulposus tissues. In addition, as a receptor that activates Wnt signaling pathway, LRP5 is mainly highly expressed in chondrocyte 5 of degenerated nucleus pulposus cells. Six genes, ANGPTL4, PTGES, IGFBP3, GDF15, TRIB3 and TNFRSF10B, which are associated with apoptosis and inflammatory responses, and are widespread in chondrocyte 4 and chondrocyte 5, may be closely related to degenerative of nucleus pulposus cells. CONCLUSIONS: Single-cell RNA sequencing revealed differential expression of Wnt/Ca2+ signaling in human normal and degenerated nucleus pulposus cells, and this differential expression may be closely related to the abundance of chondrocyte 4 and chondrocyte 5 in degenerated nucleus pulposus cells. In degenerated nucleus pulposus cells, LRP5 activate Wnt5B, which promotes nucleus pulposus cell apoptosis and inflammatory response by regulating the Wnt/Ca2+ signaling pathway, thereby promoting disc degeneration. ANGPTL4, IGFBP3, PTGES in chondrocyte 4 and TRIB3, GDF15, TNFRSF10B in chondrocyte 5 may play an important role in this process.

Research on the role and mechanism of IL-17 in intervertebral disc degeneration.

Intervertebral disc degeneration (IDD) is one of the primary causes of low back pain (LBP), which seriously affects patients' quality of life. In recent years, interleukin (IL)-17 has been shown to be highly expressed in the intervertebral disc (IVD) tissues and serum of patients with IDD, and IL-17A has been shown to promote IDD through multiple pathways. We first searched databases such as PubMed, Cochrane, Embase, and Web of Science using the search terms "IL-17 or interleukin 17″ and "intervertebral discs". The search period ranged from the inception of the databases to December 2023. A total of 24 articles were selected after full-text screening. The main conclusion of the clinical studies was that IL-17A levels are significantly increased in the IVD tissues and serum of IDD patients. The results from the in vitro studies indicated that IL-17A can activate signaling pathways such as the NF-κB and MAPK pathways; promote inflammatory responses, extracellular matrix degradation, and angiogenesis; and inhibit autophagy in nucleus pulposus cells. The main finding of the in vivo experiments was that puncture of animal IVDs resulted in elevated levels of IL-17A within the IVD, thereby inducing IDD. Clinical studies, in vitro experiments, and in vivo experiments confirmed that IL-17A is closely related to IDD. Therefore, drugs that target IL-17A may be novel treatments for IDD, providing a new theoretical basis for IDD therapy.

Ephrin B2 (EFNB2) potentially protects against intervertebral disc degeneration through inhibiting nucleus pulposus cell apoptosis.

Nucleus pulposus (NP) cell apoptosis is a significant indication of accelerated intervertebral disc degeneration; however, the precise mechanism is unelucidated as of yet. Ephrin B2 (EFNB2), the only gene down-regulated in the three degraded intervertebral disc tissue microarray groups (GSE70362, GSE147383 and GSE56081), was screened for examination in this study. Subsequently, EFNB2 was verified to be down-regulated in degraded NP tissue samples. Interleukin-1 (IL-1ß) treatment of NP cells to simulate the IDD environment indicated that IL-1ß treatment decreased EFNB2 expression. In degenerative NP cells stimulated by IL-1ß, EFNB2 knockdown significantly increased the rate of apoptosis as well as the apoptosis-related molecules cleaved-caspase-3 and the Bax to Bcl-2 ratio. EFNB2 was found to promote AKT, PI3K, and mTOR phosphorylation; the PI3K/AKT signaling role was investigated using the PI3K inhibitor LY294002. EFNB2 overexpression significantly increased PI3K/AKT pathway activity in IL-1ß-stimulated NP cells than the normal control. Moreover, EFNB2 partially alleviated NP cell apoptosis induced by IL-1ß, reduced the cleaved-cas3 level, and decreased the Bax/Bcl-2 ratio after the addition of the inhibitor LY294002. Additionally, EFNB2 overexpression inhibited the ERK1/2 phosphorylation; the effects of EFNB2 overexpression on ERK1/2 phosphorylation, degenerative NP cell viability, and cell apoptosis were partially reversed by ERK signaling activator Ceramide C6. EFNB2 comprehensively inhibited the apoptosis of NP cells by activating the PI3K/AKT signaling and inhibiting the ERK signaling, obviating the exacerbation of IDD. EFNB2 could be a potential target to protect against degenerative disc changes.

Bioenergetic dysfunction in the pathogenesis of intervertebral disc degeneration.

Intervertebral disc (IVD) degeneration is a frequent cause of low back pain and is the most common cause of disability. Treatments for symptomatic IVD degeneration, including conservative treatments such as analgesics, physical therapy, anti-inflammatories and surgeries, are aimed at alleviating neurological symptoms. However, there are no effective treatments to prevent or delay IVD degeneration. Previous studies have identified risk factors for IVD degeneration such as aging, inflammation, genetic factors, mechanical overload, nutrient deprivation and smoking, but metabolic dysfunction has not been highlighted. IVDs are the largest avascular structures in the human body and determine the hypoxic and glycolytic features of nucleus pulposus (NP) cells. Accumulating evidence has demonstrated that intracellular metabolic dysfunction is associated with IVD degeneration, but a comprehensive review is lacking. Here, by reviewing the physiological features of IVDs, pathological processes and metabolic changes associated with IVD degeneration and the functions of metabolic genes in IVDs, we highlight that glycolytic pathway and intact mitochondrial function are essential for IVD homeostasis. In degenerated NPs, glycolysis and mitochondrial function are downregulated. Boosting glycolysis such as HIF1α overexpression protects against IVD degeneration. Moreover, the correlations between metabolic diseases such as diabetes, obesity and IVD degeneration and their underlying molecular mechanisms are discussed. Hyperglycemia in diabetic diseases leads to cell senescence, the senescence-associated phenotype (SASP), apoptosis and catabolism of extracellualr matrix in IVDs. Correcting the global metabolic disorders such as insulin or GLP-1 receptor agonist administration is beneficial for diabetes associated IVD degeneration. Overall, we summarized the recent progress of investigations on metabolic contributions to IVD degeneration and provide a new perspective that correcting metabolic dysfunction may be beneficial for treating IVD degeneration.

Catalytic Nanodots-Driven Pyroptosis Suppression in Nucleus Pulposus for Antioxidant Intervention of Intervertebral Disc Degeneration.

Low back pain resulting from intervertebral disc degeneration (IVDD) is a prevalent global concern; however, its underlying mechanism remains elusive. Single-cell sequencing analyses revealed the critical involvement of pyroptosis in IVDD. Considering the involvement of reactive oxygen species (ROS) as the primary instigator of pyroptosis and the lack of an efficient intervention approach, this study developed carbonized Mn-containing nanodots (MCDs) as ROS-scavenging catalytic biomaterials to suppress pyroptosis of nucleus pulposus (NP) cells to efficiently alleviate IVDD. Catalytic MCDs have superior efficacy in scavenging intracellular ROS and rescuing homeostasis in the NP microenvironment compared with N-acetylcysteine, a classical antioxidant. The data validates that pyroptosis plays a vital role in mediating the protective effects of catalytic MCDs against oxidative stress. Systematic in vivo assessments substantiate the effectiveness of MCDs in rescuing a puncture-induced IVDD rat model, further demonstrating their ability to suppress pyroptosis. This study highlights the potential of antioxidant catalytic nanomedicine as a pyroptosis inhibitor and mechanistically unveils an efficient strategy for the treatment of IVDD.

Shared and Compartment-Specific Processes in Nucleus Pulposus and Annulus Fibrosus During Intervertebral Disc Degeneration.

Elucidating how cell populations promote onset and progression of intervertebral disc degeneration (IDD) has the potential to enable more precise therapeutic targeting of cells and mechanisms. Single-cell RNA-sequencing (scRNA-seq) is performed on surgically separated annulus fibrosus (AF) (19,978; 26,983 cells) and nucleus pulposus (NP) (20,884; 24,489 cells) from healthy and diseased human intervertebral discs (IVD). In both tissue types, depletion of cell subsets involved in maintenance of healthy IVD is observed, specifically the immature cell subsets - fibroblast progenitors and stem cells - indicative of an impairment of normal tissue self-renewal. Tissue-specific changes are also identified. In NP, several fibrotic populations are increased in degenerated IVD, indicating tissue-remodeling. In degenerated AF, a novel disease-associated subset is identified, which expresses disease-promoting genes. It is associated with pathogenic biological processes and the main gene regulatory networks include thrombospondin signaling and FOXO1 transcription factor. In NP and AF cells thrombospondin protein promoted expression of genes associated with TGFß/fibrosis signaling, angiogenesis, and nervous system development. The data reveal new insights of both shared and tissue-specific changes in specific cell populations in AF and NP during IVD degeneration. These identified mechanisms and molecules are novel and more precise targets for IDD prevention and treatment.

Abnormal stress promotes intervertebral disc degeneration through WTAP/YTHDF2-dependent TIMP3 m6A modification.

Mechanical environment worsening is an important predisposing factor that accelerates intervertebral disc degeneration (IDD), but its specific regulatory mechanisms remain unclear. In this study, we reveal the molecular mechanisms of WTAP/YTHDF2-mediated m6A modification in abnormal stress-induced intervertebral disc (IVD) matrix degradation. WTAP expression in human nucleus pulposus cells was elevated under tension. Similarly, high WTAP expression was detected in severe degenerated human and rat nucleus pulposus tissues. Functionally, WTAP was found to increase the TIMP3 transcript methylation level under tension, resulting in YTHDF2 recognition, binding, and induction of its degradation. Reduction in TIMP3 caused increases in active matrix metalloproteinases, ultimately inducing extracellular matrix degradation in nucleus pulposus cells. Macroscopically, this promotes IDD. Additionally, in vitro and in vivo inhibition of WTAP expression or TIMP3 overexpression significantly increased stress resistance in the nucleus pulposus, thereby alleviating IDD. Our results show that abnormal stress disrupts IVD matrix stability through WTAP/YTHDF2-dependent TIMP3 m6A modification.

Rescuing Nucleus Pulposus Cells from ROS Toxic Microenvironment via Mitochondria-Targeted Carbon Dot-Supported Prussian Blue to Alleviate Intervertebral Disc Degeneration.

Intervertebral disc degeneration (IVDD) is invariably accompanied by excessive accumulation of reactive oxygen species (ROS), resulting in progressive deterioration of mitochondrial function and senescence in nucleus pulposus cells (NPCs). Significantly, the main ROS production site in non-immune cells is mitochondria, suggesting mitochondria is a feasible therapeutic target to reverse IVDD. Triphenylphosphine (TPP), which is known as mitochondrial-tropic ligands, is utilized to modify carbon dot-supported Prussian blue (CD-PB) to scavenge superfluous intro-cellular ROS and maintain NPCs at normal redox levels. CD-PB-TPP can effectively escape from lysosomal phagocytosis, permitting efficient mitochondrial targeting. After strikingly lessening the ROS in mitochondria via exerting antioxidant enzyme-like activities, such as superoxide dismutase, and catalase, CD-PB-TPP rescues damaged mitochondrial function and NPCs from senescence, catabolism, and inflammatory reaction in vitro. Imaging evaluation and tissue morphology assessment in vivo suggest that disc height index, mean grey values of nucleus pulposus tissue, and histological morphology are significantly improved in the IVDD model after CD-PB-TPP is locally performed. In conclusion, this study demonstrates that ROS-induced mitochondrial dysfunction and senescence of NPCs leads to IVDD and the CD-PB-TPP possesses enormous potential to rescue this pathological process through efficient removal of ROS via targeting mitochondria, supplying a neoteric strategy for IVDD treatment.

DNMT3a-mediated methylation of PPARγ promote intervertebral disc degeneration by regulating the NF-κB pathway.

Intervertebral disc degeneration (IVDD) is a common chronic musculoskeletal disease that causes chronic low back pain and imposes an immense financial strain on patients. The pathological mechanisms underlying IVDD have not been fully elucidated. The development of IVDD is closely associated with abnormal epigenetic changes, suggesting that IVDD progression may be controlled by epigenetic mechanisms. Consequently, this study aimed to investigate the role of epigenetic regulation, including DNA methyltransferase 3a (DNMT3a)-mediated methylation and peroxisome proliferator-activated receptor γ (PPARγ) inhibition, in IVDD development. The expression of DNMT3a and PPARγ in early and late IVDD of nucleus pulposus (NP) tissues was detected using immunohistochemistry and western blotting analyses. Cellularly, DNMT3a inhibition significantly inhibited IL-1ß-induced apoptosis and extracellular matrix (ECM) degradation in rat NP cells. Pretreatment with T0070907, a specific inhibitor of PPARγ, significantly reversed the anti-apoptotic and ECM degradation effects of DNMT3a inhibition. Mechanistically, DNMT3a modified PPARγ promoter hypermethylation to activate the nuclear factor-κB (NF-κB) pathway. DNMT3a inhibition alleviated IVDD progression. Conclusively, the results of this study show that DNMT3a activates the NF-κB pathway by modifying PPARγ promoter hypermethylation to promote apoptosis and ECM degradation. Therefore, we believe that the ability of DNMT3a to mediate the PPARγ/NF-κB axis may provide new ideas for the potential pathogenesis of IVDD and may become an attractive target for the treatment of IVDD.

Mesenchymal stem cell exosomes inhibit nucleus pulposus cell apoptosis via the miR-125b-5p/TRAF6/NF-κB pathway axis.

Intervertebral disc degeneration (IVDD) is the pathological basis of a range of degenerative spinal diseases and is the primary cause of lower back pain. Mesenchymal stem cell (MSC) transplantation inhibits IVDD progression. However, the specific mechanisms that underlie these effects remain unclear. In this study, candidate microRNAs (miRNAs) are screened using bioinformatics and high-throughput sequencing. TNF-α is used to induce nucleus pulposus cell (NPC) degeneration. MSC-derived exosomes (MSC-exosomes) are obtained using high-speed centrifugation and identified by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA) and western blot analysis. Cell viability is determined by CCK-8 assay. Flow cytometry and TUNEL assays are used to detect cell apoptosis. The expression levels of miR-125b-5p are detected by RT-qPCR, and a dual-luciferase gene reporter assay confirms the downstream target genes of miR-125b-5p. Protein expression is determined by western blot analysis. Rat models are used to validate the function of miR-125b-5p in MSC-exosomes. The results show that miR-125b-5p is expressed at low levels in degenerated disc tissues compared with that in normal disc tissues; however, it is highly expressed in MSC-exosomes. Furthermore, MSC-exosomes are efficiently taken up by NPCs while miR-125b-5p is delivered into NPCs; thus, MSC-exosomes act as inhibitors of apoptosis in NPCs. Overexpression of miR-125b-5p downregulates TRAF6 expression and inhibits NF-κB activation. However, TRAF6 overexpression reverses these effects of miR-125b-5p. We demonstrate that MSC-exosomes attenuate IVDD in vivo by delivering miR-125b-5p. MSC-exosomes can deliver miR-125b-5p to target TRAF6, inhibit NF-κB activation, and attenuate the progression of IVDD.

Nucleus Pulposus-Targeting Nanocarriers Facilitate Mirna-Based Therapeutics for Intervertebral Disc Degeneration.

Intervertebral disc degeneration (IDD) is a common cause of low back pain. Understanding its molecular mechanisms is the basis for developing specific treatment. To demonstrate that miR-22-3p is critical in the regulation of IDD, miRNA microarray analyses are conducted in conjunction with in vivo and in vitro experiments. The miR-22-3p knockout (KO) mice show a marked decrease in the histological scores. Bioinformatic analysis reveals that miR-22-3p plays a mechanistic role in the development of IDD by targeting SIRT1, which in turn activates the JAK1/STAT3 signaling pathway. This is confirmed by a luciferase reporter assay and western blot analysis. Therapeutically, the delivery of miR-22-3p inhibitors and mimics through the synthesized nanoparticles in the IDD model alleviates and aggravates IDD, respectively. The nanocarriers enhance transportation of miR-22-3p to nucleus pulposus cells, thus enabling the in vivo inhibition of miR-22-3p for therapeutic purposes and consequently promoting the development of miRNA-specific drugs for IDD.

Metabolic Biomarkers Differentiate a Surgical Intervertebral Disc from a Nonsurgical Intervertebral Disc.

BACKGROUND: Degeneration of the intervertebral disc (IVD) is caused by disturbances in metabolic processes, which lead to structural disorders. The aim of this report is to analyze metabolic disorders in the degeneration process by comparing control discs with degenerated discs. In our research on the nucleus pulposus (NP), we used NMR spectroscopy of extracts of hydrophilic and hydrophobic compounds of the tissue. METHODS: Nuclear magnetic resonance (NMR) spectroscopy allows the study of biochemistry and cellular metabolism in vitro. Hydrophilic and hydrophobic compounds were extracted from the NP of the intervertebral disc. In the NMR spectra, metabolites were identified and quantitatively analyzed. The results of our research indicate disturbances in the biosynthesis and metabolism of cholesterol, the biosynthesis and degradation of various fatty acid groups, ketone bodies, or lysine, and the metabolism of glycerophospholipids, purines, glycine, inositol, galactose, alanine, glutamate, and pyruvate in the biosynthesis of valine and isoleucine, leucine. All these disorders indicate pathomechanisms related to oxidative stress, energy, neurotransmission disturbances, and disturbances in the structure and functioning of cell membranes, inflammation, or chronic pain generators. CONCLUSIONS: NMR spectroscopy allows the identification of metabolites differentiating surgical from nonsurgical discs. These data may provide guidance in in vivo MRS studies in assessing the severity of lesions of the disc.

Circ-STC2 promotes the ferroptosis of nucleus pulposus cells via targeting miR-486-3p/TFR2 axis.

BACKGROUND: Low back pain (LBP) has become the second leading cause of disability worldwide, which has brought great economic burden to people. It is generally believed that intervertebral disc degeneration (IDD) is the main cause of LBP. This study aimed to explore the role of circ-STC2 in the pathogenesis of IDD. METHODS: Nucleus pulposus cells (NPCs) were treated with T-Butyl Hydrogen Peroxide (TBHP) to establish IDD model in vitro. RT-qPCR was performed to detect mRNA expressions. The cell viability was detected with CCK-8 assay. The levels of lactate dehydrogenase (LDH), malondialdehyde (MDA), Fe2+ and glutathione (GSH) of NPCs were measured by corresponding kits. The protein expressions were determined by western blot. Dual-luciferase reporter and RNA pull-down assays were conducted to verify the relationship between circ-STC2 or transferrin recepto 2 (TFR2) and miR-486-3p. RESULTS: Circ-STC2 and TFR2 expressions were up-regulated in IDD tissues, and miR-486-3p expression was down-regulated. Knockdown of circ-STC2 promoted the cell viability and inhibited the ferroptosis of the NPCs. The GSH levels, and glutathione peroxidase 4 (GPX4) and solute carrier family 7 member 11 (SLC7A11) protein expressions were increased, the LDH, MDA and Fe2+ levels and achaete-scute complexlike 4 (ASCL4) protein expressions were decreased after circ-STC2 knockdown. Knockdown of miR-486-3p abrogated the si-circ-STC2 effects and overexpression of TFR2 reversed the miR-486-3p mimic effects. CONCLUSIONS: Circ-STC2 inhibits the cell viability, induced the ferroptosis of the TBHP treated NPCs via targeting miR-486-3p/TFR2 axis.

Roles of pyroptosis in intervertebral disc degeneration.

Intervertebral disc degeneration (IDD), the key pathological process in low back pain, is characterized by chronic inflammation and progressive cell death. Pyroptosis is a type of pro-inflammatory programmed necrosis mediated by inflammasomes that is dependent on the gasdermin family of proteins. An in-depth study of the pathological mechanisms of IDD has revealed that pyroptosis plays an important role in its occurrence and development. The molecular characteristics and activation signaling mechanisms of pyroptosis are reviewed in this paper. Moreover, the specific roles of pyroptosis in IDD pathology are outlined and various targeted drugs for its treatment are highlighted.

A Redox Homeostasis Modulatory Hydrogel with GLRX3+ Extracellular Vesicles Attenuates Disc Degeneration by Suppressing Nucleus Pulposus Cell Senescence.

Characterized by nucleus pulposus (NP) cell senescence and extracellular matrix (ECM) degradation, disc degeneration is a common pathology for various degenerative spinal disorders. To date, effective treatments for disc degeneration are absent. Here, we found that Glutaredoxin3 (GLRX3) is an important redox-regulating molecule associated with NP cell senescence and disc degeneration. Using a hypoxic preconditioning method, we developed GLRX3+ mesenchymal stem cell-derived extracellular vehicles (EVs-GLRX3), which enhanced the cellular antioxidant defense, thus preventing reactive oxygen species (ROS) accumulation and senescence cascade expansion in vitro. Further, a disc tissue-like biopolymer-based supramolecular hydrogel, which was injectable, degradable, and ROS-responsive, was proposed to deliver EVs-GLRX3 for treating disc degeneration. Using a rat model of disc degeneration, we demonstrated that the EVs-GLRX3-loaded hydrogel attenuated mitochondrial damage, alleviated the NP senescence state, and restored ECM deposition by modulating the redox homeostasis. Our findings suggested that modulation of redox homeostasis in the disc can rejuvenate NP cell senescence and thus attenuate disc degeneration.