Effect of platelet-rich fibrin matrix in complex with artificial material Nubiplant on expression of chondrogenic marker genes and morphogenesis of the nucleus pulposus cells of intervertebral discs in rats

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).

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

Oxidative Medicine and Cellular Longevity ◽

10.1155/2021/4657014 ◽

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.

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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

International Journal of Polymer Science ◽

10.1155/2019/8925807 ◽

2019 ◽

Vol 2019 ◽

pp. 1-7 ◽

Cited By ~ 2

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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Transformation of resident notochord‐descendent nucleus pulposus cells in mouse injury‐induced fibrotic intervertebral discs

Aging Cell ◽

10.1111/acel.13254 ◽

2020 ◽

Vol 19 (11) ◽

Cited By ~ 1

Author(s):

Tiffany Y. K. Au ◽

To‐Kam Lam ◽

Yan Peng ◽

Sarah L. Wynn ◽

Kenneth M. C. Cheung ◽

...

Keyword(s):

Nucleus Pulposus ◽

Intervertebral Discs ◽

Nucleus Pulposus Cells

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A20 of nucleus pulposus cells plays a self-protection role via the nuclear factor-kappa B pathway in the inflammatory microenvironment

Bone and Joint Research ◽

10.1302/2046-3758.95.bjr-2019-0230.r1 ◽

2020 ◽

Vol 9 (5) ◽

pp. 225-235

Author(s):

Xin Peng ◽

Cong Zhang ◽

Jun-Ping Bao ◽

Lei Zhu ◽

Rui Shi ◽

...

Keyword(s):

Nucleus Pulposus ◽

Messenger Rna ◽

Nuclear Factor Kappa B ◽

Intervertebral Discs ◽

Control Group ◽

Nuclear Factor ◽

Nucleus Pulposus Cells ◽

Necrosis Factor Alpha ◽

Nuclear Factor Kappa ◽

Tnf Α

Aims Inflammatory response plays a pivotal role in the pathophysiological process of intervertebral disc degeneration (IDD). A20 (also known as tumour necrosis factor alpha-induced protein 3 (TNFAIP3)) is a ubiquitin-editing enzyme that restricts nuclear factor-kappa B (NF-κB) signalling. A20 prevents the occurrence of multiple inflammatory diseases. However, the role of A20 in the initiation of IDD has not been elucidated. The aim of the study was to investigate the effect of A20 in senescence of TNF alpha (TNF-α)-induced nucleus pulposus cells (NPCs). Methods Immunohistochemical staining was performed to observe the expression of A20 in normal and degenerated human intervertebral discs. The NPCs were dissected from the tail vertebrae of healthy male Sprague-Dawley rats and were cultured in the incubator. In the experiment, TNF-α was used to mimic the inflammatory environment of IDD. The cell viability and senescence were examined to investigate the effect of A20 on TNF-α-treated NPCs. The expression of messenger RNA (mRNA)-encoding proteins related to matrix macromolecules (collagen II, aggrecan) and senescence markers (p53, p16). Additionally, NF-κB/p65 activity of NPCs was detected within different test compounds. Results The expression of A20 was upregulated in degenerate human intervertebral discs. The A20 levels of NPCs in TNF-α inflammatory microenvironments were dramatically higher than those of the control group. TNF-α significantly decreased cell proliferation potency but increased senescence-associated beta-galactosidase (SA-β-Gal) activity, the expression of senescence-associated proteins, the synthesis of extracellular matrix, and G1 cycle arrest. The senescence indicators and NF-κB/p65 expression of A20 downregulated group treated with TNF-α were significantly upregulated compared to TNF-α-treated normal NPCs. Conclusion A20 has a self-protective effect on the senescence of NPCs induced by TNF-α. The downregulation of A20 in NPCs exacerbated the senescence of NPCs induced by TNF-α. Cite this article: Bone Joint Res. 2020;9(5):225–235.

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Mechanobiology of annulus fibrosus and nucleus pulposus cells in intervertebral discs

Cell and Tissue Research ◽

10.1007/s00441-019-03136-1 ◽

2019 ◽

Vol 379 (3) ◽

pp. 429-444 ◽

Cited By ~ 2

Author(s):

Sara Molladavoodi ◽

John McMorran ◽

Diane Gregory

Keyword(s):

Nucleus Pulposus ◽

Annulus Fibrosus ◽

Intervertebral Discs ◽

Nucleus Pulposus Cells

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Identification of critical genes in nucleus pulposus cells isolated from degenerated intervertebral discs using bioinformatics analysis

Molecular Medicine Reports ◽

10.3892/mmr.2017.6662 ◽

2017 ◽

Vol 16 (1) ◽

pp. 553-564 ◽

Cited By ~ 6

Author(s):

Zhuangchen Zhu ◽

Guang Chen ◽

Wei Jiao ◽

Defeng Wang ◽

Yan Cao ◽

...

Keyword(s):

Nucleus Pulposus ◽

Bioinformatics Analysis ◽

Intervertebral Discs ◽

Nucleus Pulposus Cells ◽

Degenerated Intervertebral Discs

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Mangiferin Alleviates Mitochondrial ROS in Nucleus Pulposus Cells and Protects against Intervertebral Disc Degeneration via Suppression of NF-κB Signaling Pathway

Oxidative Medicine and Cellular Longevity ◽

10.1155/2021/6632786 ◽

2021 ◽

Vol 2021 ◽

pp. 1-27

Author(s):

Haichao Yu ◽

Guowei Hou ◽

Jiankang Cao ◽

Yanyu Yin ◽

Yunpeng Zhao ◽

...

Keyword(s):

Intervertebral Disc ◽

Mitochondrial Dysfunction ◽

Signaling Pathway ◽

Nucleus Pulposus ◽

Disc Degeneration ◽

Intervertebral Disc Degeneration ◽

Intervertebral Discs ◽

Nucleus Pulposus Cells ◽

Mitochondrial Ros ◽

Tnf Α

Intervertebral disc degeneration (IVDD), one of the most common clinical diseases worldwide, causes disc herniation and sciatica. Recent studies have identified the involvement of mitochondrial dysfunction, inflammatory responses, and extracellular matrix degradation in IVDD. Mangiferin is known to protect against various diseases by inhibiting oxidative stress, suppressing inflammation reaction, and relieving mitochondrial dysfunction. Whether mangiferin can alleviate IVDD remains to be elucidated. In the present study, human nucleus pulposus cells (HNPCs) and mouse intervertebral discs were cultured and stimulated with TNF-α, with or without treatment of mangiferin. Moreover, we established a rat needle puncture model and injected mangiferin into the intervertebral discs to verify its protective effect on IVDD. Furthermore, the activity of the NF-κB signaling pathway was tested in vitro. Our results indicated that mangiferin alleviated the inflammatory response and reversed the loss of major intervertebral disc components. Besides, mangiferin reduced reactive oxygen species production, ameliorated mitochondrial damage, and decreased the expression of apoptosis-related parameters in stimulation of TNF-α. In addition, mangiferin antagonized the activation of the NF-κB signaling pathway induced by TNF-α. Collectively, mangiferin antagonized mitochondrial ROS in NP cells and protected against IVDD by suppressing the activation of the NF-κB signaling pathway, which might provide a potential therapeutic instrument for IVDD.

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Is Osteogenic Differentiation of Human Nucleus Pulposus Cells a Possibility for Biological Spinal Fusion?

Cartilage ◽

10.1177/1947603518754628 ◽

2018 ◽

Vol 11 (2) ◽

pp. 181-191 ◽

Cited By ~ 3

Author(s):

Sharon J. Brown ◽

Sarah A. Turner ◽

Birender S. Balain ◽

Neil T. Davidson ◽

Sally Roberts

Keyword(s):

Extracellular Matrix ◽

Osteogenic Differentiation ◽

Spinal Fusion ◽

Nucleus Pulposus ◽

Monolayer Culture ◽

Intervertebral Discs ◽

Nucleus Pulposus Cells ◽

Alizarin Red ◽

Dihydroxyvitamin D ◽

Polymerase Chain

Objective The purpose of this study was to investigate whether a simple, biologically robust method for inducing calcification of degenerate intervertebral discs (IVD) could be developed to provide an alternative treatment for patients requiring spinal fusion. Design Nucleus pulposus (NP) cells isolated from 14 human IVDs were cultured in monolayer and exposed to osteogenic medium, 1,25-dihydroxyvitamin D3 (VitD3), parathyroid hormone (PTH), and bone morphogenic proteins (BMPs) 2/7 to determine if they could become osteogenic. Similarly explant cultures of IVDs from 11 patients were cultured in osteogenic media with and without prior exposure to VitD3 and BMP-2. Osteogenic differentiation was assessed by alkaline phosphatase activity and areas of calcification identified by alizarin red or von Kossa staining. Expression of osteogenic genes during monolayer culture was determined using polymerase chain reaction and explant tissues assessed for BMP inhibitors. Human bone marrow–derived mesenchymal stromal cells (MSCs) were used for comparison. Results Standard osteogenic media was optimum for promoting mineralization by human NP cells in monolayer. Some osteogenic differentiation was observed with 10 nM VitD3, but none following application of PTH or BMPs. Regions of calcification were detected in 2 of the eleven IVD tissue explants, one cultured in osteogenic media and one with the addition of VitD3 and BMP-2. Conclusions Human NP cells can become osteogenic in monolayer and calcification of the extracellular matrix can also occur, although not consistently. Inhibitory factors within either the cells or the extracellular matrix may hinder osteogenesis, indicating that a robust biological fusion at this time requires further optimization.

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