Background: Radiculitis-induced pain (RIP) results from dorsal root ganglion (DRG) sensitization due to inflammation. Hypoxia-inducible factor 1-alpha (HIF-1α) is linked to inflammatory responses through metabolic changes, but its role in RIP is not well understood. Gua Sha therapy has been shown to reduce inflammation and neural damage from lumbar disc herniation (LDH). This study investigates whether HIF-1α-mediated metabolic reprogramming contributes to the pain-relieving effects of Gua Sha in RIP. Methods: Male SD rats were subjected to LDH surgery and divided into six groups: sham, model, sham Gua Sha, Gua Sha, Gua Sha + DMOG, and Gua Sha + YC-1. Gua Sha was applied 5 days postsurgery, every other day for three sessions per course, totaling three courses. Changes in paw withdrawal threshold (PWT) and latency (PWL) were monitored, along with blood flow in the rats' backs. Levels of IL-1β, TNF-α, and NF-κB were assessed in serum and DRG tissue. Pathological changes and hypoxia in DRG tissues were observed using hematoxylin-eosin staining and immunofluorescence. Western blotting and qPCR measured HIF-1α, GLUT1, PFKM, and PDK1 expression, while lactic acid and ATP levels in DRG tissue were also evaluated. Results: Gua Sha increased PWT and PWL, reduced serum and DRG inflammatory factors, improved back microcirculation, alleviated DRG hypoxia, and decreased HIF-1α and related signaling factors. It also lowered lactic acid and raised ATP levels. DMOG, a HIF-1α activator, reversed these effects. HIF-1α activation did not affect serum inflammatory factors but partially improved PWT. Inhibition of HIF-1α with YC-1 did not significantly differ from Gua Sha alone. Conclusion: HIF-1α-mediated metabolic reprogramming is a pathogenic mechanism in RIP. Gua Sha alleviates RIP by enhancing microcirculation, improving DRG hypoxia, inhibiting HIF-1α-mediated reprogramming, and reducing DRG sensitization and inflammation. This study provides insights into the mechanisms of Gua Sha's therapeutic effects in RIP.

Traditional methods of plasmid delivery, including viral vectors, lipofection, and electroporation, are widely used for gene editing but have limitations, such as cellular toxicity, limited transfection efficiency in primary cells, and nonspecific side effects. Here, we report the development of nucleus pulposus cell (NPC)-mimicking nanoparticles (HIF1A@NNP) with an NPC membrane as the shell and pcDNA3.1+-rHIF1A encapsulated in the core via extrusion. HIF1A@NNP exhibited a protein expression pattern similar to that of the NPC membrane and displayed a typical vesicle profile. Compared to liposomes and lentiviruses, HIF1A@NNP overexpressed HIF1A in NPCs while improving cell viability. HIF1A@NNP was more readily internalized by NPCs than by other cell types, with fewer effects on vascularization, nerve growth, and macrophage polarization than HIF1A overexpression using lipo3000. HIF1A@NNP reduced the apoptotic rate and inhibited the senescent phenotype, as evidenced by reduced DNA damage, lower levels of senescence-related proteins, and fewer SA-β-Gal-positive cells. HIF1A@NNP induced a senescence-associated secretory phenotype (SASP), which enhanced macrophage migration and M1 polarization. Additionally, HIF1A@NNP activated autophagy in NPCs. In summary, HIF1A@NNP demonstrated satisfactory biocompatibility, alleviated the SASP, and inhibited SASP-mediated macrophage recruitment and inflammatory polarization, leading to reduced disc degeneration and providing a promising strategy for combating intervertebral disc degeneration. STATEMENT OF SIGNIFICANCE: Conventional plasmid delivery methods like viral vectors, lipofection, and electroporation struggle with cellular toxicity and inefficiency in primary cells. Non-cell-specific HIF1A activation via these methods may exacerbate inflammation and pain, as HIF1A drives angiogenesis and dendritic ingrowth into the disc. Thus, a cell-specific delivery strategy could circumvent such adverse effects. Our study introduces HIF1A@NNP, a nanoparticle mimicking nucleus pulposus cells (NPCs), with an NPC membrane shell encapsulating pcDNA3.1+-rHIF1A. It preferentially targets NPCs, achieving superior HIF1A overexpression and cell viability compared to liposomes and lentiviruses. This represents a highly promising and potentially transformative approach against intervertebral disc degeneration.

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.

A comprehensive understanding of the molecules that play key roles in the physiological and pathological homeostasis of the human intervertebral disc (IVD) remains challenging, as does the development of new therapeutic treatments. We recently found a positive correlation between IVD degeneration (IDD) and P2X7 receptor (P2X7R) expression increases both in the cytoplasm and in the nucleus. Using immunocytochemistry, reverse transcription PCR (RT-PCR), overexpression, and chromatin immunoprecipitation, we found that NFATc1 and hypoxia-inducible factor-1α (HIF-1α) are critical regulators of P2X7R. Both transcription factors are recruited at the promoter of the P2RX7 gene and involved in its positive and negative regulation, respectively. Furthermore, using the proximity ligation assay, we revealed that P2X7R and NFATc1 form a molecular complex and that P2X7R is closely associated with lamin A/C, a major component of the nuclear lamina. Collectively, our study identifies, for the first time, P2X7R and NFATc1 as markers of IVD degeneration and demonstrates that both NFATc1 and lamin A/C are interaction partners of P2X7R.

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.

Accumulation of advanced glycation end products (AGEs) causes apoptosis in human nucleus pulposus cells (NPCs), contributing to intervertebral disc degeneration (IVDD). The purpose of this study was to determine the roles of thioredoxin-interacting protein (TXNIP) in the mechanisms underlying AGE-induced apoptosis of NPCs. TXNIP was silenced or overexpressed in HNPCs exposed to AGEs. Glycolysis was assessed using extracellular acidification rate (ECAR), ATP level, GLUT1, and GLUT4 measurements. AGEs, TXNIP, GLUT1, and GLUT4 levels in IVDD patients were measured as well. In NPCs, AGEs reduced cell viability, induced apoptosis, inhibited glycolysis, and increased TXNIP expression. Silencing TXNIP compromised the effects of AGEs on cell viability, apoptosis, and glycolysis in NPCs. Furthermore, TXNIP overexpression resulted in decreased cell viability, increased apoptotic cells, and glycolysis suppression. Furthermore, co-treatment with a glycolysis inhibitor improved TXNIP silencing's suppressive effects on AGE-induced cell injury in NPCs. In IVDD patients with Pfirrmann Grades II-V, increasing trends in AGEs and TXNIP were observed, while decreasing trends in GLUT1 and GLUT4. AGE levels had positive correlations with TXNIP levels. Both AGE and TXNIP levels correlated negatively with GLUT1 and GLUT4. Our study indicates that TXNIP plays a role in mediating AGE-induced cell injury through suppressing glycolysis. The accumulation of AGEs, the upregulation of TXNIP, and the downregulation of GLUT1 and GLUT4 are all linked to the progression of IVDD.

The dysregulation of intracellular and extracellular environments as well as the aberrant expression of ion channels on the cell membrane are intricately linked to a diverse array of degenerative disorders, including intervertebral disc degeneration. This condition is a significant contributor to low back pain, which poses a substantial burden on both personal quality of life and societal economics. Changes in the number and function of ion channels can disrupt the water and ion balance both inside and outside cells, thereby impacting the physiological functions of tissues and organs. Therefore, maintaining ion homeostasis and stable expression of ion channels within the cellular microenvironment may prove beneficial in the treatment of disc degeneration. Aquaporin (AQP), calcium ion channels, and acid-sensitive ion channels (ASIC) play crucial roles in regulating water, calcium ions, and hydrogen ions levels. These channels have significant effects on physiological and pathological processes such as cellular aging, inflammatory response, stromal decomposition, endoplasmic reticulum stress, and accumulation of cell metabolites. Additionally, Piezo 1, transient receptor potential vanilloid type 4 (TRPV4), tension response enhancer binding protein (TonEBP), potassium ions, zinc ions, and tungsten all play a role in the process of intervertebral disc degeneration. This review endeavors to elucidate alterations in the microenvironment of the nucleus pulposus during intervertebral disc degeneration (IVDD), with a view to offer novel insights and approaches for exploring therapeutic interventions against disc degeneration.

Hypoxia can promote stem cell proliferation and migration through HIF-1α. Hypoxia can regulate cellular endoplasmic reticulum (ER) stress. Some studies have reported the relationship among hypoxia, HIF-α, and ER stress, however, while little is known about HIF-α and ER stress in ADSCs under hypoxic conditions. The purpose of the study was to investigate the role and relationship of hypoxic conditions, HIF-1α and ER stress in regulating adipose mesenchymal stem cells (ADSCs) proliferation, migration, and NPC-like differentiation.

ADSCs were pretreated with hypoxia, HIF-1α gene transfection, and HIF-1α gene silence. The ADSCs proliferation, migration, and NPC-like differentiation were assessed. The expression of HIF-1α in ADSCs was regulated; then, the changes of ER stress level in ADSCs were observed to investigate the relationship between ER stress and HIF-1α in ADSCs under hypoxic conditions.

The cell proliferation and migration assay results show that hypoxia and HIF-1α overexpression can significantly increase the ADSCs proliferation and migration, while HIF-1α inhibition can significantly decrease the ADSCs proliferation and migration. The HIF-1α and co-cultured with NPCs played an important role in the directional differentiation of ADSCs into NPCs. The hypoxia-regulated ER stress in ADSCs through the HIF-1α pathway, thereby regulating the cellular state of ADSCs, was also observed.

Hypoxia and HIF-1α play important roles in proliferation, migration, and NPC-like differentiation of ADSCs. This study provides preliminary evidence that HIF-1α-regulated ER stress thus affects ADSCs proliferation, migration, and differentiation. Therefore, HIF-1α and ER may serve as key points to improve the efficacy of ADSCs in treating disc degeneration.

The intervertebral disc (IVD) aids in motion and acts to absorb energy transmitted to the spine. With little inherent regenerative capacity, degeneration of the intervertebral disc results in intervertebral disc disease, which contributes to low back pain and significant disability in many individuals. Increasing evidence suggests that IVD degeneration is a disease of the whole joint that is associated with significant inflammation. Moreover, studies show elevated macrophage accumulation within the IVD with increasing levels of disease severity; however, we still need to understand the roles, be they causative or consequential, of macrophages during the degenerative process. In this narrative review, we discuss hallmarks of IVD degeneration, showcase evidence of macrophage involvement during disc degeneration, and explore burgeoning research aimed at understanding the molecular pathways regulating macrophage functions during intervertebral disc degeneration.

The degenerative disease of the intervertebral disc is nowadays an important health problem, which has still not been understood and solved adequately. The vertebral endplate is regarded as one of the vital elements in the structure of the intervertebral disc. Its constituent cells, the chondrocytes in the endplate, may also be involved in the process of the intervertebral disc degeneration and their role is central both under physiological and pathological conditions. They main functions include a role in homeostasis of the extracellular environment of the intervertebral disc, metabolic support and nutrition of the discal nucleus and annulus beneath and the preservation of the extracellular matrix. Therefore, it is understandable that the cells in the endplate have been in the centre of research from several viewpoints, such as development, degeneration and growth, reparation and remodelling, as well as treatment strategies. In this article, we briefly review the importance of vertebral endplate, which are often overlooked, in the intervertebral disc degeneration.

Cellular niches play fundamental roles in regulating cellular behaviors. However, the effect of niches on direct converted cells remains unexplored. In the present study, the specific combination of transcription factors is first identified to directly acquire induced nucleus pulposus-like cells (iNPLCs). Next, tunable physical properties of collagen niches are fabricated based on various crosslinking degrees. Collagen niches significantly affect actomyosin cytoskeleton and then influence the maturation of iNPLCs. Using gain- and loss of function approaches, the appropriate physical states of collagen niches are found to significantly enhance the maturation of iNPLCs through actomyosin contractility. Moreover, in a rat model of degenerative disc diseases, iNPLCs with collagen niches are transplanted into the lesion to achieve significant improvements. As a result, overexpression of transcription factors in human dermal fibroblasts are efficiently converted into iNPLCs and the optimal collagen niches affect cellular cytoskeleton and then facilitate iNPLCs maturation toward human nucleus pulposus cells. These findings encourage more in-depth studies toward the interactions of niches and direct conversion, which would contribute to the development of direct conversion.

Despite extensive efforts over the past 40 years, there is still a significant gap in knowledge of the characteristics of elastic fibers in the intervertebral disc (IVD). More studies are required to clarify the potential contribution of elastic fibers to the IVD (healthy and diseased) function and recommend critical areas for future investigations. On the other hand, current IVD in-vitro models are not true reflections of the complex biological IVD tissue and the role of elastic fibers has often been ignored in developing relevant tissue-engineered scaffolds and realistic computational models. This has affected the progress of IVD studies (tissue engineering solutions, biomechanics, fundamental biology) and translation into clinical practice. Motivated by the current gap, the current review paper presents a comprehensive study (from the early 1980s to 2022) that explores the current understanding of structural (multi-scale hierarchy), biological (development and aging, elastin content, and cell-fiber interaction), and biomechanical properties of the IVD elastic fibers, and provides new insights into future investigations in this domain.