scholarly journals Propionibacterium acnes Accelerates Intervertebral Disc Degeneration by Inducing Pyroptosis of Nucleus Pulposus Cells via the ROS-NLRP3 Pathway

2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Guoqing Tang ◽  
Xiaoguang Han ◽  
Zhijie Lin ◽  
Hongbin Qian ◽  
Bing Chen ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
Propionibacterium Acnes ◽  
Strong Association ◽  
Intervertebral Discs ◽  
Inflammatory Factors ◽  
Death Process ◽  
Dependent Manner ◽  
Nucleus Pulposus Cells ◽  
Alternative Treatment Strategy ◽  
Ivd Degeneration

Our previous study verified the occurrence of Propionibacterium acnes (P. acnes), a low-virulence anaerobic bacterium, latently residing in degenerated intervertebral discs (IVDs), and the infection had a strong association with IVD degeneration. We explored whether P. acnes induces nucleus pulposus cell (NPC) pyroptosis, a more dangerous cell death process than apoptosis, and accelerates IVD degeneration via the pyroptotic products interleukin- (IL-) 1β and IL-18. After coculturing with P. acnes, human NPCs showed significant upregulation of NOD-like receptor 3 (NLRP3), cleaved IL-1β, cleaved caspase-1, and cleaved gasdermin D in response to the overexpression of IL-1β and IL-18 in a time- and dose-dependent manner. In addition, the gene expression of inflammatory factors and catabolic enzymes significantly increased in normal NPCs when cocultured with pyroptotic NPCs in a transwell system, and the adverse effects were inhibited when NPC pyroptosis was suppressed. Furthermore, inoculation of P. acnes into the IVDs of rats caused significant pyroptosis of NPCs and remarkable IVD degeneration. Finally, coculture of NPCs with P. acnes induced the overexpression of reactive oxygen species (ROS) and NLRP3, while inhibition of both factors reduced NPC pyroptosis. Therefore, P. acnes induces NPC pyroptosis via the ROS-NLRP3 signaling pathway, and the pyroptotic NPCs cause an IVD degeneration cascade. Targeting the P. acnes-induced pyroptosis of NPCs may become an alternative treatment strategy for IVD degeneration in the future.

scholarly journals Spatially defined single-cell transcriptional profiling characterizes diverse chondrocyte subtypes and nucleus pulposus progenitors in human intervertebral discs

Bone Research ◽  
2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Yibo Gan ◽  
Jian He ◽  
Jun Zhu ◽  
Zhengyang Xu ◽  
Zhong Wang ◽  
...  
Keyword(s):  
Single Cell ◽  
Nucleus Pulposus ◽  
Molecular Mechanisms ◽  
Transcriptional Profiling ◽  
Intervertebral Discs ◽  
Cellular Heterogeneity ◽  
Effector Functions ◽  
Cartilage Endplate ◽  
Intervertebral Disks ◽  
Ivd Degeneration

AbstractA comprehensive understanding of the cellular heterogeneity and molecular mechanisms underlying the development, homeostasis, and disease of human intervertebral disks (IVDs) remains challenging. Here, the transcriptomic landscape of 108 108 IVD cells was mapped using single-cell RNA sequencing of three main compartments from young and adult healthy IVDs, including the nucleus pulposus (NP), annulus fibrosus, and cartilage endplate (CEP). The chondrocyte subclusters were classified based on their potential regulatory, homeostatic, and effector functions in extracellular matrix (ECM) homeostasis. Notably, in the NP, a PROCR+ resident progenitor population showed enriched colony-forming unit-fibroblast (CFU-F) activity and trilineage differentiation capacity. Finally, intercellular crosstalk based on signaling network analysis uncovered that the PDGF and TGF-β cascades are important cues in the NP microenvironment. In conclusion, a single-cell transcriptomic atlas that resolves spatially regulated cellular heterogeneity together with the critical signaling that underlies homeostasis will help to establish new therapeutic strategies for IVD degeneration in the clinic.

scholarly journals Non-viral reprogramming of human nucleus pulposus cells with FOXF1 via extracellular vesicle delivery: an in vitro and in vivo study

2021 ◽  
Vol 41 ◽  
pp. 90-107
Author(s):  
S Tang ◽  
◽  
A Salazar-Puerta ◽  
J Richards ◽  
S Khan ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
Viral Vectors ◽  
Extracellular Vesicle ◽  
Proteoglycan Synthesis ◽  
Nucleus Pulposus Cells ◽  
Keratin 19 ◽  
Altered Gene ◽  
Ivd Degeneration

Intervertebral disc (IVD) degeneration is characterized by decreased cellularity and proteoglycan synthesis and increased inflammation, catabolism, and neural/vascular ingrowth. Regenerative methods for IVD degeneration are largely cell-therapy-based or involve viral vectors, which are associated with mutagenesis and undesired immune responses. The present study used bulk electroporation and engineered extracellular vesicles (EVs) to deliver forkhead-box F1 (FOXF1) mRNA to degenerate human nucleus pulposus (NP) cells as a minimally invasive therapeutic strategy for IVD regeneration. Bulk electroporation was used to investigate FOXF1 effects on human NP cells during a 4-week culture in 3D agarose constructs. Engineered EV delivery of FOXF1 into human IVD cells in monolayer was determined, with subsequent in vivo validation in a pilot mouse IVD puncture model. FOXF1 transfection significantly altered gene expression by upregulating healthy NP markers [FOXF1, keratin 19 (KRT19)], decreasing inflammatory cytokines [interleukin (IL)-1β, -6], catabolic enzymes [metalloproteinase 13 (MMP13)] and nerve growth factor (NGF), with significant increases in glycosaminoglycan accumulation in human NP cells. Engineered EVs loaded with FOXF1 demonstrated successful encapsulation of FOXF1 cargo and effective uptake by human NP cells cultured in monolayer. Injection of FOXF1-loaded EVs into the mouse IVD in vivo resulted in a significant upregulation of FOXF1 and Brachyury, compared to controls at 7 d post-injection, with no evidence of cytotoxicity. This is the first study to demonstrate non-viral delivery of FOXF1 and reprogramming of human NP cells in vitro and mouse IVD cells in vivo. This strategy represents a non-addictive approach for treating IVD degeneration and associated back pain.

scholarly journals The REDD1/TXNIP Complex Accelerates Oxidative Stress-Induced Apoptosis of Nucleus Pulposus Cells through the Mitochondrial Pathway

2021 ◽  
Vol 2021 ◽  
pp. 1-22
Author(s):  
Huipeng Yin ◽  
Kun Wang ◽  
Abhirup Das ◽  
Gaocai Li ◽  
Yu Song ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Intervertebral Disc ◽  
Nucleus Pulposus ◽  
Cell Apoptosis ◽  
Redox Homeostasis ◽  
Mitochondrial Pathway ◽  
Nucleus Pulposus Cells ◽  
Damage Response ◽  
Induced Apoptosis ◽  
Ivd Degeneration

The death of nucleus pulposus (NP) cells is an important cause of intervertebral disc (IVD) degeneration. Redox disturbance caused by dysfunctional mitochondria has been considered as a vital risk for NP cell survival. It is valuable to identify key proteins maintaining mitochondrial function in NP cells. A previous study found that regulated in development and DNA damage response 1 (REDD1) are upregulated during intervertebral disc degeneration and that REDD1 can cause NP cell apoptosis. Thus, the present study further explores the effect of REDD1 on IVD degeneration. Our results showed that REDD1 promotes NP cell apoptosis via the mitochondrial pathway. Importantly, REDD1 formed a complex with TXNIP to strengthen its own action, and the combination was consolidated under H2O2-induced oxidative stress. The combined inhibition of the REDD1/TXNIP complex was better than that of REDD1 or TXNIP alone in restoring cell proliferation and accelerating apoptosis. Moreover, p53 acts as the transcription factor of REDD1 to regulate the REDD1/TXNIP complex under oxidative stress. Altogether, our results demonstrated that the REDD1/TXNIP complex mediated H2O2-induced human NP cell apoptosis and IVD degeneration through the mitochondrial pathway. Interferences on these sites to achieve mitochondrial redox homeostasis may be a novel therapeutic strategy for oxidative stress-associated IVD degeneration.

scholarly journals LIPUS inhibits inflammation and catabolism through the NF‐κB pathway in human degenerative nucleus pulposus cells

2021 ◽  
Vol 16 (1) ◽  
Author(s):  
Weiwei Yi ◽  
Qing Chen ◽  
Chuan Liu ◽  
Kaiting Li ◽  
Bailong Tao ◽  
...  
Keyword(s):  
Protein Expression ◽  
Real Time ◽  
Signaling Pathway ◽  
Nucleus Pulposus ◽  
Real Time Pcr ◽  
Inflammatory Factors ◽  
Nucleus Pulposus Cells ◽  
Tnf Α ◽  
Gene And Protein Expression ◽  
Collagen Ii

Abstract Background Low-intensity pulsed ultrasound (LIPUS) is a safe and noninvasive rehabilitative physical therapy with anti-inflammatory effects. The current study investigated the effect of LIPUS on the inflammation of nucleus pulposus (NP) cells and its underlying mechanism. Methods Human NP cells were acquired from lumbar disc herniation tissue samples and cultured for experiments. Human NP cells were treated with LPS and then exposed to LIPUS (15 mW/cm2, 30 mW/cm2 and 60 mW/cm2) for 20 min daily for 3 days to determine the appropriate intensity to inhibit the expression of the inflammatory factors TNF-α and IL-1β. The gene and protein expression of aggrecan, collagen II, MMP-3 and MMP-9 was measured by real‐time PCR and western blotting, respectively. The activity of the nuclear factor‐kappa B (NF‐κB) pathway was examined by western blotting and immunofluorescence. After pretreatment with the NF-κB inhibitor PDTC, the expression of TNF-α, IL-1β, MMP-3 and MMP-9 was measured by real‐time PCR. Results LIPUS at intensities of 15 mW/cm2, 30 mW/cm2 and 60 mW/cm2 inhibited LPS-induced NP cell expression of the inflammatory factors TNF-α and IL-1β, especially at 30 mW/cm2. LIPUS significantly upregulated the gene and protein expression of aggrecan and collagen II and downregulated the gene and protein expression of MMP-3 and MMP-9 in LPS-induced NP cells. The NF‐κB signaling pathway was inhibited by LIPUS through inhibiting the protein expression of p-P65 and the translocation of P65 into the nucleus in LPS-induced NP cells. In addition, LIPUS had similar effects as the NF-κB inhibitor PDTC by inhibiting the NF-κB signaling pathway, inflammation and catabolism in LPS-induced human degenerative nucleus pulposus cells. Conclusion LIPUS inhibited inflammation and catabolism through the NF‐κB pathway in human degenerative nucleus pulposus cells.

scholarly journals Effect of Lentivirus-mediated GDF5 Transfection on Differentiation of Rabbit Nucleus Pulposus Mesenchymal Stem Cells

2020 ◽  
Author(s):  
kun zhu ◽  
Rui Zhao ◽  
Yuchen Ye ◽  
Gang Xu ◽  
Changchun Zhang
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Nucleus Pulposus ◽  
Lentiviral Vector ◽  
Intervertebral Discs ◽  
Degenerative Disease ◽  
The Other ◽  
Control Group ◽  
Nucleus Pulposus Cells ◽  
Qrt Pcr

Abstract Background: Disc degenerative disease is a common senile degenerative disease, which seriously affects the quality of life of patients.The purpose of this study is to observe the biological and cytological characteristics of rabbit nucleus pulposus mesenchymal stem cells (NPMSCs), and to determine the effect of growth differentiation factor 5(GDF5) on the differentiation of rabbit NPMSCs by lentivirus transfection.Methods: In vitro culture model of rabbit NPMSCs was established and NPMSCs cells were identified by flow cytometry (FCM)and quantitative real-time PCR(qRT-PCR). Then NPMSCs were divided into three groups: lentiviral vector carrying GDF5 was used to transfect NPMSCs, to determine the transfection rate, which was recorded as transfection group, and the NPMSCs transfected with ordinary lentiviral vector was recorded as control group, NPMSCs without processing was recorded as normal group. FCM, qRT-PCR and Western Blot(WB) were used to detected the change of NPMSCs.Results: The transfected NPMSCs by GDF5 became longer and narrower, and the cell density decreased,and the positive rate of GDF5 in the transfected group was significantly higher than that in the other two groups (P<0.05). The mRNA expression of KRT8, KRT18, KRT19 in the transfected group was significantly higher than the other two groups(P<0.05),the result of WB were the same to qRT-PCR. Conclusions: GDF5 can induce the differentiation of NPMSCs and repair degenerative intervertebral discs. Lentiviral vector carrying GDF5 can be integrated into the chromosome genome of NPMSCs and promote differentiation of NPMSCs into nucleus pulposus cells(NPCs).

Effect of Inflammation on the Osmotic Response of Nucleus Pulposus Cells

2012 ◽  
Author(s):  
Robert Maidhof ◽  
Neena Rajan ◽  
Nadeen O. Chahine
Keyword(s):  
Nucleus Pulposus ◽  
Biomechanical Properties ◽  
Nucleus Pulposus Cells ◽  
Cellular Mechanisms ◽  
Osmotic Response ◽  
Tnf Α ◽  
Disc Cells ◽  
Cytokine Stimulation ◽  
Ivd Degeneration

Intervertebral disc (IVD) degeneration is accompanied by elevated levels of pro-inflammatory cytokines, particularly IL-1β and TNF-α [1]. Disc cells from the nucleus pulposus (NPs) respond to cytokine stimulation with increased catabolic breakdown of the tissue, resulting in a positive feedback of disc integrity loss and further inflammation [2]. Previous studies by our group have examined the response of NP cells to Toll-Like Receptor-4 (TLR-4) activation through stimulation with lipopolysaccharide (LPS). TLR-4 is a pattern recognition receptor that is activated in innate immunity and by polysaccharide fragments from degenerated proteoglycans. TLR-4 activation by LPS results in stimulation of multiple cytokines by NP cells [3]. Moreover, we have shown that in vivo LPS injection results in catabolic changes in the IVD, including matrix breakdown, decrease in biomechanical properties and loss of disc height [4]. However, the specific cellular mechanisms for these catabolic changes remain to be elucidated.

scholarly journals Regenerative Response of Degenerate Human Nucleus Pulposus Cells to GDF6 Stimulation

2020 ◽  
Vol 21 (19) ◽  
pp. 7143
Author(s):  
Tom Hodgkinson ◽  
Hamish T. J. Gilbert ◽  
Tej Pandya ◽  
Ashish D. Diwan ◽  
Judith A. Hoyland ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
Annulus Fibrosus ◽  
Therapeutic Strategies ◽  
Signalling Pathways ◽  
Signal Transduction Pathways ◽  
Enzyme Expression ◽  
Nucleus Pulposus Cells ◽  
Ivd Degeneration ◽  
Homeostatic Mechanisms ◽  
Catabolic Enzyme

Growth differentiation factor (GDF) family members have been implicated in the development and maintenance of healthy nucleus pulposus (NP) tissue, making them promising therapeutic candidates for treatment of intervertebral disc (IVD) degeneration and associated back pain. GDF6 has been shown to promote discogenic differentiation of mesenchymal stem cells, but its effect on NP cells remains largely unknown. Our aim was to investigate GDF6 signalling in adult human NP cells derived from degenerate tissue and determine the signal transduction pathways critical for GDF6-mediated phenotypic changes and tissue homeostatic mechanisms. This study demonstrates maintained expression of GDF6 receptors in human NP and annulus fibrosus (AF) cells across a range of degeneration grades at gene and protein level. We observed an anabolic response in NP cells treated with recombinant GDF6 (increased expression of matrix and NP-phenotypic markers; increased glycosaminoglycan production; no change in catabolic enzyme expression), and identified the signalling pathways involved in these responses (SMAD1/5/8 and ERK1/2 phosphorylation, validated by blocking studies). These findings suggest that GDF6 promotes a healthy disc tissue phenotype in degenerate NP cells through SMAD-dependent and -independent (ERK1/2) mechanisms, which is important for development of GDF6 therapeutic strategies for treatment of degenerate discs.

scholarly journals Protective Effect of Polygonatum sibiricum Polysaccharides on Apoptosis, Inflammation, and Oxidative Stress in Nucleus Pulposus Cells of Rats with the Degeneration of the Intervertebral Disc

2019 ◽  
Vol 2019 ◽  
pp. 1-7 ◽  
Author(s):  
Zhaohui Zhai ◽  
Zhaoxin Li ◽  
Zhonglei Ji ◽  
Xiaosheng Lu
Keyword(s):  
Oxidative Stress ◽  
Intervertebral Disc ◽  
Nucleus Pulposus ◽  
Disc Degeneration ◽  
Rat Model ◽  
Intervertebral Disc Degeneration ◽  
Intervertebral Discs ◽  
New Drugs ◽  
Nucleus Pulposus Cells ◽  
And Oxidative Stress

Objective. Polygonatum sibiricum polysaccharide (PSP) has antioxidant activity, immune enhancement, and other biological properties. However, the effect of PSP on intervertebral disc degeneration has not been reported. In this study, we mainly investigated the effect of PSP on the apoptosis, inflammation, and oxidative stress of nucleus pulposus cells (NPCs) during the process of intervertebral disc degeneration. Methods. A rat NPC model induced by H2O2 was constructed. The CCK8 method was used to measure the effects of PSP on the apoptosis of rat NPCs induced by H2O2. The effects on the activity of SOD and content of MDA were also determined. The rat model of intervertebral disc degeneration was treated with PSP for 1 month, and the mRNA expression levels of IL-1β, COX2, iNOS, Col2α1, Col10α1, and MMP3 were measured by qPCR in the tissue of intervertebral disc. NPCs from the degenerated intervertebral discs were separated, and the cell viability was measured by the CCK8 method. The contents of SOD and MDA in NPCs were determined as well. Results. PSP significantly reduced the apoptosis of NPCs induced by H2O2, significantly increased the SOD content, and decreased the content of MDA in H2O2-induced NPCs. The expression level of IL-1β, COX2, and iNOS in the rat model with intervertebral disc degeneration was significantly downregulated after 1 month of PSP treatment. PSP treatment increased the expression of Col2α1 type and significantly decreased the expression of Col10α1 type collagen and MMP3 in rats with disc degeneration. PSP treatment significantly reduced NPC apoptosis and increased its SOD content and reduced MDA content, which is consistent with the results from cell-level experiments. Conclusion. PSP can effectively reduce the apoptosis, inflammation, and oxidative stress of H2O2-induced NPCs in rats with intervertebral disc degeneration and mitigate the progression of intervertebral disc degeneration, which has the potential to be developed as new drugs for the treatment of intervertebral disc degeneration.

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