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 Knockdown of miR-660 protects nucleus pulposus cells from TNF-a-induced apoptosis by targeting serum amyloid A1

2020 ◽  
Vol 15 (1) ◽  
Author(s):  
Hao Jie Zhang ◽  
Xue Hai Ma ◽  
Song Lin Xie ◽  
Shu lian Qin ◽  
Cong Zhi Liu ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
General Characteristic ◽  
Luciferase Reporter ◽  
Healthy Controls ◽  
Nucleus Pulposus Cells ◽  
Apoptosis Rate ◽  
Tnf Α ◽  
Significant Difference ◽  
Induced Apoptosis

Abstract Background Intervertebral disc degeneration (IVDD) is a well-known cause of lower back pain, which is induced by multiple factors including increased apoptosis and decreased survival of nucleus pulposus cells. In this study, we evaluate the effect and potential mechanism of miR-660 on the nucleus pulposus cells apoptosis induced by TNF-α. Methods First, we collected tissue of nucleus pulposus from IVDD and healthy controls. General characteristic of the IVDD and healthy control was also collected. And, we also collected nucleus pulposus cells that stimulated by TNF-α or control. miRNA microarray was performed to identify the differentially expressed miRNAs. Apoptosis rate and miR-660 relative expression was measured after stimulated with different concentration of TNF-α to identify the optimal concentration of TNF-α. Second, we successfully constructed antigomiR-660 to block the miR-660 expression in nucleus pulposus cells and then stimulated with TNF-α (100 ng/ml, 12 h). The apoptosis rates and relative protein expression were then measured again. The target association between miR-660 and SAA1 was confirmed by dual-luciferase reporter. Results There was no significant difference between the age (IVDD: 39 ± 10 years, healthy controls: 36 ± 7 years), BMI and sex between IVDD and healthy controls. Microarray analysis found that miR-660 was significantly up-regulated in IVDD and TNF-α treated groups, which was further identified by PCR. We found that the rate of apoptosis and miR-660 expression increased with TNF-α concentration increased. Finally, TNF-a with 100 ng/ml was used for further experiment. Compared with TNF-α group, TNF-α + antigomiR-660 could significantly down-regulated the apoptosis rate and relative protein (c-Caspase3 and c-Caspase7). Dual-luciferase reporter revealed that miR-660 could directly binding to the SAA1 at 80–87 sites. Compared with TNF-α alone group, TNF-α + antigomiR-660 significantly up-regulated the SAA1 expression (P < 0.05). Conclusion These results indicated that knockdown of miR-660 protected the nucleus pulposus from apoptosis that induced TNF-α via up-regulation of SAA1. Further studies should focus on the role of miR-660 in protecting IVDD in vivo.

scholarly journals LRP5-deficiency in OsxCreERT2 mice models intervertebral disc degeneration by aging and compression

2019 ◽  
Author(s):  
Matthew J. Silva ◽  
Nilsson Holguin
Keyword(s):  
Transcription Factor ◽  
Intervertebral Disc ◽  
Wnt Signaling ◽  
Cell Fate ◽  
Biomechanical Properties ◽  
Nucleus Pulposus Cells ◽  
Hypertrophic Chondrocyte ◽  
Age Related ◽  
Ivd Degeneration

ABSTRACTOsterix is a critical transcription factor of mesenchymal stem cell fate, where its loss or loss of WNT signaling diverts differentiation to a chondrocytic lineage. Intervertebral disc (IVD) degeneration activates differentiation of prehypertrophic chondrocyte-like cells and inactivates WNT signaling, but its interacting role with osterix is unclear. First, compared to young-adult (5mo), mechanical compression of old (18mo) IVD induced greater IVD degeneration. Aging (5 vs 12mo) and/or compression reduced the transcription of osterix and notochordal marker T by 40-75%. Compression elevated transcription of hypertrophic chondrocyte marker MMP13 and pre-osterix transcription factor RUNX2, but less so in 12mo IVD. Next, using an Ai9/td reporter and immunohistochemistry, annulus fibrosus and nucleus pulposus cells of 5mo IVD expressed osterix, but aging and compression reduced its expression. Lastly, in vivo LRP5-deficiency in osterix-expressing cells degenerated the IVD, inactivated WNT signaling, reduced the biomechanical properties by 45-70%, and reduced transcription of osterix, notochordal markers and chondrocytic markers by 60-80%. Overall, these data indicate that age-related inactivation of WNT signaling in osterix-expressing cells may limit regeneration by depleting progenitors and attenuating the expansion of chondrocyte-like cells.

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.

In Vivo Administration of an Inflammatory Stimulant can Trigger Loss of Biomechanical and Biochemical Properties of the Intervertebral Disc

2012 ◽  
Author(s):  
Neena Rajan ◽  
Nate Stetson ◽  
Robert Maidhof ◽  
Mitchell Levine ◽  
Nadeen Chahine
Keyword(s):  
Intervertebral Disc ◽  
Morphological Changes ◽  
Direct Injection ◽  
Biomechanical Properties ◽  
Biochemical Properties ◽  
Post Injury ◽  
Sham Injection ◽  
Tnf Α ◽  
Ivd Degeneration

Human intervertebral disc (IVD) degeneration is accompanied by chronic inflammation, particularly seen in the elevated levels of pro-inflammatory cytokines IL-1β and TNF-α [1–3]. Animal models of disc degeneration (DD) using stab or laceration of the disc generally reproduce morphological changes of IVD degeneration. However, inflammatory changes in these models are thought to be acute and transient post injury [4–6]. The goal of this study is to explore the effect of direct inflammatory stimulation of the IVD on disc biochemical and biomechanical properties in vivo. We utilize lipopolysaccharide (LPS), an inflammatory stimulant that provokes secretion of multiple cytokines by disc cells. We have previously shown that direct injection of LPS into the disc results in significantly higher protein levels of IL-1β, TNF-α, HMGB-1 and MIF vs. sham injection up to 7 days post administration [7]. The goal of this study is to explore the dose-dependent response of this inflammatory stimulation on the biochemical and biomechanical properties of IVD in vivo. We hypothesize that LPS stimulation mimics the pathophysiology of DD by triggering a group of cytokines that are associated with IVD degeneration. LPS is administered using micro needles (<10% disc height) in order to minimize the potential disruption by needle injection.

scholarly journals CB2-mediated attenuation of nucleus pulposus degeneration via the amelioration of inflammation and oxidative stress in vivo and in vitro

2021 ◽  
Vol 27 (1) ◽  
Author(s):  
Xiaoqiang Cheng ◽  
Jiayi Lin ◽  
Zhanghuan Chen ◽  
Yubo Mao ◽  
Xiexin Wu ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Nucleus Pulposus ◽  
Nucleus Pulposus Cell ◽  
Immunocytochemical Staining ◽  
Nucleus Pulposus Cells ◽  
Collagen Ii ◽  
Ivd Degeneration ◽  
Oxide Synthase

Abstract Background Nucleus pulposus cell (NPC) degeneration is widely accepted as one of the major causes of intervertebral disc (IVD) degeneration (IVDD). The pathogenesis of IVDD is complex and consists of inflammation, oxidative stress, and the loss of extracellular matrix (ECM). Cannabinoid type 2 receptor (CB2) has been shown to be involved in the pathological mechanism of a variety of diseases due to its anti-inflammatory effects and antioxidative stress capacity. Method In Vitro, H2O2 was used to induce degeneration of nucleus pulposus cells, mRNA and protein expression level was determined by RT-PCR and Western Blot, and Immunocytochemical staining were used to detect expression of collagen II, aggrecan, MMP3/13, superoxide dismutase 2 (SOD2) and inducible nitric oxide synthase (iNOS). In vivo, the potential therapeutic effect of CB2 was detected in the rat acupuncture model. Result In vitro, we found that the CB2 agonist (JWH133) treatment reduced the oxidative stress level in NPCs induced by hydrogen peroxide (H2O2) treatment. Furthermore, the expression of inflammatory cytokines was also decreased by JWH133 treatment. We found that collagen II and aggrecan expression was preserved, whereas matrix metalloproteinase levels were reduced. In vivo, we established a rat model by needle puncture. Imaging assessment revealed that the disc height index (DHI) and morphology of IVD were significantly improved, and the disc degeneration process was delayed by treatment of JWH133. Furthermore, immunohistochemical (IHC) staining revealed that JWH133 could inhibit the degradation of collagen II and decrease the expression of MMP3. Conclusions The experiment indicates the oxidative stress and inflammatory response of rat NPCs induced by H2O2 could be inhibited by activating CB2. This study reveals that CB2 activation can effectively delay the development of IVDD, providing an effective therapeutic target for IVDD.

scholarly journals Retinoic acid inhibits the pyroptosis of degenerated nucleus pulposus cells by activating Sirt1-SOD2 signaling

2021 ◽  
Author(s):  
Peng-Fei Li ◽  
Fei Xiong ◽  
Ying Yin ◽  
Hong-Yuan Xing ◽  
Shao-Jun Hu ◽  
...  
Keyword(s):  
Retinoic Acid ◽  
Intervertebral Disc ◽  
Nucleus Pulposus ◽  
Underlying Mechanism ◽  
Ros Generation ◽  
Common Disease ◽  
Nucleus Pulposus Cells ◽  
Ivd Degeneration ◽  
Related Proteins

Abstract Background: Intervertebral disc (IVD) degeneration is a common disease and initiated by the degeneration of nucleus pulposus (NP). The pyroptosis of degenerated NP cells (dNPCs) plays an important role in NP degeneration and may be a potential target in the treatment of IVD degeneration. The purpose of this study is to identify a feasible solution that can inhibit NP cell pyroptosis to therapy the degeneration of the intervertebral disc. Result: In this study, we determined the effects of retinoic acid (RA) on dNPCs and investigated the underlying mechanism of RA mediated pyroptosis in dNPCs. We also verified the effects of RA on IVD degeneration in vivo. Our results demonstrated that RA significantly increased the proliferation and the protein expression of sox9, aggrecan, and collagen II of dNPCs. Pyroptosis-related proteins such as cleaved caspase-1, NT-GSDMD, IL-1β, IL-18, and the pyroptosis rate of dNPCs was significantly decreased by RA. We also found that Sirt1-SOD2 signaling was activated, while ROS generation and TXNIP/NLRP3 signaling in dNPCs was inhibited after the addition of RA. Furthermore, RA also recovered the structure of NP and increased the contents of sGAG and collagen in vivo. Conclusion Our study demonstrated that RA can inhibit the pyroptosis and increase the ECM synthesis function of dNPCs and verified that RA has a protective effect in IVD degeneration.

Compartmentalization of the matrix formed by nucleus pulposus and annulus fibrosus cells in alginate gel

2002 ◽  
Vol 30 (6) ◽  
pp. 874-878 ◽  
Author(s):  
E. Thonar ◽  
H. An ◽  
K. Masuda
Keyword(s):  
Nucleus Pulposus ◽  
Annulus Fibrosus ◽  
Articular Chondrocyte ◽  
Alginate Gel ◽  
Age Related ◽  
The Matrix ◽  
Disc Cells ◽  
A Cell ◽  
Associated Matrix

Intervertebral disc cells cultured in alginate gel are capable of reforming in alginate, a matrix that consists of two compartments: a rim of metabolically active cell-associated matrix and a more abundant, but metabolically less active, further removed matrix. At any one age and in most species, the cell-associated matrix formed by a nucleus pulposus or annulus fibrosus cell cultured in this way is less abundant than that formed by an articular chondrocyte. In both the cell-associated matrix and further removed matrix, the ratio of aggrecan to collagen is significantly higher in the case of nucleus pulposus than of annulus fibrosus, a feature that also distinguishes the matrices of the nucleus pulposus and annulus fibrosus in vivo. Nucleus pulposus and annulus fibrosus cells from older donors show a decreased ability to reform a cell-associated matrix rich in aggrecan. There is, however, some evidence that gene therapy and/or exposure of the cells to defined stimulatory factors can help overcome some of these age-related limitations. This contention is supported by recent evidence that nucleus pulposus and annulus fibrosus cells from adult donors can be manipulated to form, using the recently developed alginate-recovered chondrocyte system, a resilient tissue that bears many of the characteristics of the tissue in which these cells reside in vivo.

MicroRNA-155 suppresses the catabolic effect induced by TNF-α and IL-1β by targeting C/EBPβ in rat nucleus pulposus cells

2018 ◽  
Vol 60 (2) ◽  
pp. 165-177 ◽  
Author(s):  
Jie Zhou ◽  
Anjing Liang ◽  
Junmin Hong ◽  
Jianchao Sun ◽  
Xiaolin Lin ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
Nucleus Pulposus Cells ◽  
Tnf Α

scholarly journals Injectable Hydrogel Combined with Nucleus Pulposus-Derived Mesenchymal Stem Cells for the Treatment of Degenerative Intervertebral Disc in Rats

2019 ◽  
Vol 2019 ◽  
pp. 1-17 ◽  
Author(s):  
Feng Wang ◽  
Li-ping Nan ◽  
Shi-feng Zhou ◽  
Yang Liu ◽  
Ze-yu Wang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Intervertebral Disc ◽  
Nucleus Pulposus ◽  
Rat Model ◽  
Injectable Hydrogel ◽  
Cell Scaffold ◽  
Ivd Degeneration

Stem cell-based tissue engineering in treating intervertebral disc (IVD) degeneration is promising. An appropriate cell scaffold can maintain the viability and function of transplanted cells. Injectable hydrogel has the potential to be an appropriate cell scaffold as it can mimic the condition of the natural extracellular matrix (ECM) of nucleus pulposus (NP) and provide binding sites for cells. This study was aimed at investigating the effect of injectable hydrogel-loaded NP-derived mesenchymal stem cells (NPMSC) for the treatment of IVD degeneration (IDD) in rats. In this study, we selected injectable 3D-RGD peptide-modified polysaccharide hydrogel as a cell transplantation scaffold. In vitro, the biocompatibility, microstructure, and induced differentiation effect on NPMSC of the hydrogel were studied. In vivo, the regenerative effect of hydrogel-loaded NPMSC on degenerated NP in a rat model was evaluated. The results showed that NPMSC was biocompatible and able to induce differentiation in hydrogel in vivo. The disc height index (almost 87%) and MRI index (3313.83±227.79) of the hydrogel-loaded NPMSC group were significantly higher than those of other groups at 8 weeks after injection. Histological staining and immunofluorescence showed that the hydrogel-loaded NPMSC also partly restored the structure and ECM content of degenerated NP after 8 weeks. Moreover, the hydrogel could support long-term NPMSC survival and decrease cell apoptosis rate of the rat IVD. In conclusion, injectable hydrogel-loaded NPMSC transplantation can delay the level of IDD and promote the regeneration of the degenerative IVD in the rat model.

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