scholarly journals N-Cadherin-Mediated Activation of PI3K/Akt-GSK-3β Signaling Attenuates Nucleus Pulposus Cell Apoptosis Under High-Magnitude Compression

2017 ◽  
Vol 44 (1) ◽  
pp. 229-239 ◽  
Author(s):  
Pei Li ◽  
Zherui Liang ◽  
Gang Hou ◽  
Lei Song ◽  
Ruijie Zhang ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
Cell Apoptosis ◽  
Caspase 3 ◽  
Nucleus Pulposus Cell ◽  
Underlying Mechanism ◽  
Akt Pathway ◽  
Cell Phenotype ◽  
High Magnitude ◽  
Gsk 3Β

Background/Aims: Mechanical overloading-induced nucleus pulposus (NP) apoptosis plays an important role in the pathogenesis of intervertebral disc degeneration. N-cadherin (N-CDH)-mediated signaling preserves normal NP cell phenotype. This study aims to investigate the effects of N-CDH on NP cell apoptosis under high-magnitude compression and the underlying mechanism behind this process. Methods: Rat NP cells seeded on scaffold were perfusion-cultured using a self-developed perfusion bioreactor for 5 days and experienced different magnitudes (2% and 20% compressive deformation, respectively) of compression at a frequency of 1.0 Hz for 4 hours once per day. The un-loaded NP cells were used as controls. Lentivirus-mediated N-CDH overexpression and inhibitor LY294002 were used to further investigate the role of N-CDH and PI3K/Akt pathway under high-magnitude compression, respectively. NP cell apoptosis was evaluated by caspase-3 activity measured using a commercial kit, flow cytometry, and expression of apoptosis-related molecules analyzed by real-time PCR and western blotting assays. Results: High-magnitude compression significantly increased apoptotic NP cells, caspase-3 activity and expression of pro-apoptotic molecules (Bax and caspase-3/cleaved caspase-3), but decreased expression of anti-apoptotic molecule (Bcl-2). High-magnitude compression decreased expression of N-CDH, p-Akt and p-GSK-3β. However, N-CDH overexpression attenuated NP cell apoptosis and increased expression of p-Akt and p-GSK-3β under high-magnitude compression. Further analysis showed that inhibition of the PI3K/Akt pathway suppressed NP cell apoptosis and decreased expression of p-GSK-3β, but had no significant effects on N-CDH expression under high-magnitude compression. Conclusion: N-CDH can attenuate NP cell apoptosis through activating the PI3K/Akt-GSK-3β signaling under high-magnitude compression.

scholarly journals Melatonin Suppresses Apoptosis of Nucleus Pulposus Cells through Inhibiting Autophagy via the PI3K/Akt Pathway in a High-Glucose Culture

2021 ◽  
Vol 2021 ◽  
pp. 1-8
Author(s):  
Jian Li ◽  
Chengqiang Wang ◽  
Lixin Xue ◽  
Fan Zhang ◽  
Jianqiang Liu
Keyword(s):  
Protein Expression ◽  
Nucleus Pulposus ◽  
Disc Degeneration ◽  
High Glucose ◽  
Cell Apoptosis ◽  
Caspase 3 ◽  
Akt Pathway ◽  
Protective Effects ◽  
Nucleus Pulposus Cells

Diabetes mellitus- (DM-) associated hyperglycemia promotes apoptosis of disc nucleus pulposus (NP) cells, which is a contributor to intervertebral disc degeneration (IDD). Melatonin is able to protect against cell apoptosis. However, its effects on apoptosis of NP cell in a high-glucose culture remain unclear. The purpose of the present study was to investigate the effects and molecular mechanism of melatonin on NP cell apoptosis in a high-glucose culture. NP cells were cultured in the baseline medium supplemented with a high-glucose concentration (0.2 M) for 3 days. The control cells were only cultured in the baseline medium. Additionally, the pharmaceutical inhibitor LY294002 was added along with the culture medium to investigate the possible role of the PI3K/Akt pathway. Apoptosis, autophagy, and activity of the PI3K/Akt pathway of NP cells among these groups were evaluated. Compared with the control NP cells, high glucose significantly increased cell apoptosis ratio and caspase-3/caspase-9 activity and decreased mRNA expression of Bcl-2, whereas it increased mRNA or protein expression of Bax, caspase-3, cleaved caspase-3, cleaved PARP, and autophagy-related molecules (Atg3, Atg5, Beclin-1, and LC3-II) and decreased protein expression of p-Akt compared with the control cells. Additionally, melatonin partly inhibited the effects of high glucose on those parameters of cell apoptosis, autophagy, and activation of PI3K/Akt. In conclusion, melatonin attenuates apoptosis of NP cells through inhibiting the excessive autophagy via the PI3K/Akt pathway in a high-glucose culture. This study provides new theoretical basis of the protective effects of melatonin against disc degeneration in a DM patient.

scholarly journals Resveratrol inhibits IL-1β-mediated nucleus pulposus cell apoptosis through regulating the PI3K/Akt pathway

2019 ◽  
Vol 39 (3) ◽  
Author(s):  
Yanhai Jiang ◽  
Zhijie Xie ◽  
Jinying Yu ◽  
Lianqiang Fu
Keyword(s):  
Nucleus Pulposus ◽  
Disc Degeneration ◽  
Cell Apoptosis ◽  
Inflammatory Cytokine ◽  
Culture Medium ◽  
Caspase 3 ◽  
Signal Transduction Pathway ◽  
Akt Pathway ◽  
Protective Effects ◽  
Apoptosis Ratio

Abstract Nucleus pulposus (NP) cell apoptosis is a classical cellular character during intervertebral disc degeneration (IDD). Previous studies have shown that inflammatory cytokine-induced NP cell apoptosis plays an important role in disc degeneration. The present study was aimed to investigate whether resveratrol can suppress IL-1β-mediated NP cell apoptosis and the potential signal transduction pathway. Experimental rat NP cells were treated with culture medium containing IL-1β (20 ng/ml) for 7 days. Control NP cells were cultured in the baseline medium. Resveratrol was added along with culture medium to investigate its effects. The inhibitor LY294002 was used to study the role of the PI3K/Akt pathway. NP cell apoptosis was reflected by the caspase-3 activity, cell apoptosis ratio, and expression of apoptosis-related molecules (Bcl-2, Bax, caspase-3, cleaved caspase-3, and cleaved PARP). Compared with the control NP cells, IL-1β significantly increased caspase-3 activity, NP cell apoptosis ratio and mRNA/protein expression of Bax, caspase-3, cleaved caspase-3 and cleaved PARP, but decreased mRNA expression of Bcl-2. However, resveratrol partly suppressed the effects of IL-1β on those cell apoptosis-related parameters. Further analysis showed that IL-1β significantly decreased activity of the PI3K/Akt pathway whereas resveratrol partly increased activity of the PI3K/Akt pathway in NP cells treated with IL-1β. Additionally, when the inhibitor LY294002 was added along with the resveratrol, its protective effects against IL-1β-induced NP cell apoptosis were attenuated. In conclusion, resveratrol suppresses IL-1β-mediated NP cell apoptosis through activating the PI3K/Akt pathway. Resveratrol may be an effective drug to attenuate inflammatory cytokine-induced disc degenerative changes.

scholarly journals N-Cadherin Maintains the Healthy Biology of Nucleus Pulposus Cells under High-Magnitude Compression

2017 ◽  
Vol 43 (6) ◽  
pp. 2327-2337 ◽  
Author(s):  
Zhenyu Wang ◽  
Jiali Leng ◽  
Yuguang Zhao ◽  
Dehai Yu ◽  
Feng Xu ◽  
...  
Keyword(s):  
Cell Viability ◽  
Nucleus Pulposus ◽  
Cell Biology ◽  
Mechanical Load ◽  
Cell Phenotype ◽  
Nucleus Pulposus Cells ◽  
Keratin 19 ◽  
High Magnitude ◽  
And Control

Background/Aims: Mechanical load can regulate disc nucleus pulposus (NP) biology in terms of cell viability, matrix homeostasis and cell phenotype. N-cadherin (N-CDH) is a molecular marker of NP cells. This study investigated the role of N-CDH in maintaining NP cell phenotype, NP matrix synthesis and NP cell viability under high-magnitude compression. Methods: Rat NP cells seeded on scaffolds were perfusion-cultured using a self-developed perfusion bioreactor for 5 days. NP cell biology in terms of cell apoptosis, matrix biosynthesis and cell phenotype was studied after the cells were subjected to different compressive magnitudes (low- and high-magnitudes: 2% and 20% compressive deformation, respectively). Non-loaded NP cells were used as controls. Lentivirus-mediated N-CDH overexpression was used to further investigate the role of N-CDH under high-magnitude compression. Results: The 20% deformation compression condition significantly decreased N-CDH expression compared with the 2% deformation compression and control conditions. Meanwhile, 20% deformation compression increased the number of apoptotic NP cells, up-regulated the expression of Bax and cleaved-caspase-3 and down-regulated the expression of Bcl-2, matrix macromolecules (aggrecan and collagen II) and NP cell markers (glypican-3, CAXII and keratin-19) compared with 2% deformation compression. Additionally, N-CDH overexpression attenuated the effects of 20% deformation compression on NP cell biology in relation to the designated parameters. Conclusion: N-CDH helps to restore the cell viability, matrix biosynthesis and cellular phenotype of NP cells under high-magnitude compression.

Role of the ERK1/2 pathway in osmolarity effects on nucleus pulposus cell apoptosis in a disc perfusion culture

2016 ◽  
Vol 35 (1) ◽  
pp. 86-92 ◽  
Author(s):  
Pei Li ◽  
Yibo Gan ◽  
Haoming Wang ◽  
Yuan Xu ◽  
Songtao Li ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
Cell Apoptosis ◽  
Perfusion Culture ◽  
Nucleus Pulposus Cell

scholarly journals Osteogenic protein-1 attenuates apoptosis and enhances matrix synthesis of nucleus pulposus cells under high magnitude compression though inhibiting the p38 MAPK pathway

2018 ◽  
Vol 38 (2) ◽  
Author(s):  
Haolin Fang ◽  
Xianzhou Li ◽  
Haiming Shen ◽  
Buwei Sun ◽  
Haijun Teng ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
P38 Mapk ◽  
Disc Degeneration ◽  
Cell Apoptosis ◽  
Caspase 3 ◽  
Mapk Pathway ◽  
High Magnitude ◽  
P38 Mapk Pathway ◽  
Regulated Expression ◽  
Osteogenic Protein

Disc degeneration is correlated with mechanical load. Osteogenic protein-1 (OP-1) is potential to regenerate degenerative disc. To investigate whether OP-1 can protect against high magitude compression-induced nucleus pulposus (NP) cell apoptosis and NP matrix catabolism, and its potential mechanism; porcine discs were cultured in a bioreactor and compressed at a relatively high-magnitude mechanical compression (1.3 MPa at a frequency of 1.0 Hz for 2 h once per day) for 7 days. OP-1 was added along with the culture medium to investigate the protective effects of OP-1. NP cell apoptosis and matrix biosynthesis were evaluated. Additionally, activity of the p38 MAPK pathway is also analyzed. Compared with the control group, high magnitude compression significantly promoted NP cell apoptosis and decreased NP matrix biosynthesis, reflected by the increase in the number of terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL)-positive cells and caspase-3 activity, the up-regulated expression of Bax and caspase-3 mRNA and down-regulated expression of Bcl-2 mRNA, and the decreased Alcian Blue staining intensity and expression of matrix proteins (aggrecan and collagen II). However, OP-1 addition partly attenuated the effects of high magnitude compression on NP cell apoptosis and NP matrix biosynthesis. Further analysis showed that inhibition of the p38 MAPK pathway partly participated in this process. OP-1 can attenuate high magnitude compression-induced NP cell apoptosis and promoted NP matrix biosynthesis, and inhibition of the p38 MAPK pathway may participate in this regulatory process. The present study provides that OP-1 may be efficient in retarding mechanical overloading-exacerbated disc degeneration.

scholarly journals Resveratrol attenuates high glucose-induced nucleus pulposus cell apoptosis and senescence through activating the ROS-mediated PI3K/Akt pathway

2018 ◽  
Vol 38 (2) ◽  
Author(s):  
Wenping Wang ◽  
Pei Li ◽  
Jiagang Xu ◽  
Xiangkun Wu ◽  
Zhiliang Guo ◽  
...  
Keyword(s):  
Nucleus Pulposus ◽  
High Glucose ◽  
Cell Apoptosis ◽  
Culture Medium ◽  
Nucleus Pulposus Cell ◽  
Akt Pathway ◽  
Control Group ◽  
Ros Generation ◽  
Protective Effects ◽  
Potential Mechanism

Background: Diabetes mellitus is closely correlated with disc degeneration. Nucleus pulposus (NP) cell apoptosis and senescence are typical cellular features within the degenerative disc. Resveratrol is a newly identified phytoalexin that has protective effects on cartilaginous tissue. Objective: To investigate the whether resveratrol can protect against high glucose-induced NP cell apoptosis and senescence, and the potential mechanism in this process. Methods: Rat NP cells were cultured in either 10% FBS culture medium (control group) or 10% FBS with a high glucose concentration (0.2 M, experiment group) for 3 days. Resveratrol or the combination of resveratrol and LY294002 was added into the culture medium of experiment group to investigate the effects of resveratrol and the PI3K/Akt pathway. Results: High glucose significantly promoted NP cell apoptosis and NP cell senescence compared with the control group. Resveratrol exhibited protective effects against high glucose-induced NP cell apoptosis and senescence. Further analysis showed that resveratrol suppressed reactive oxygen species (ROS) generation and increased the activity of the PI3K/Akt pathway under the high glucose condition. However, the LY294002 had no significant effects on ROS content in the resveratrol-treated high glucose group. Conclusion: Resveratrol can attenuate high glucose-induced NP cell apoptosis and senescence, and the activation of ROS-mediated PI3K/Akt pathway may be the potential mechanism in this process.

scholarly journals Osteogenic protein-1 attenuates nucleus pulposus cell apoptosis through activating the PI3K/Akt/mTOR pathway in a hyperosmotic culture

2018 ◽  
Vol 38 (6) ◽  
Author(s):  
Yan Yang ◽  
Xiyang Wang ◽  
Zheng Liu ◽  
Xiao Xiao ◽  
Wenkai Hu ◽  
...  
Keyword(s):  
Protein Expression ◽  
Nucleus Pulposus ◽  
Cell Apoptosis ◽  
Caspase 3 ◽  
Mtor Pathway ◽  
Protective Effects ◽  
Fk 506 ◽  
Osteogenic Protein ◽  
Apoptosis Ratio

Background: Previous studies have indicated that osteogenic protein-1 has protective effects on the biological functions of intervertebral disc cells. Hyperosmolarity is an important physicochemical factor within the disc nucleus pulposus (NP) region, which obviously promotes NP cell apoptosis. Objective: To study the effects of osteogenic protein-1 (OP-1) on NP cell apoptosis induced by hyperosmolarity and the potential signaling transduction pathway. Methods: Rat NP cells were cultured in a hyperosmotic medium with or without OP-1 addition for 7 days. Inhibitor 294002 and inhibitor FK-506 were used to investigate the role of the PI3K/Akt/mTOR pathway in this process. NP cell apoptosis were evaluated by cell apoptosis ratio, activity of caspase-3/9 and gene/protein expression of apoptosis-related molecules (Bax, Bcl-2, caspase-3/cleaved caspase-3 and cleaved PARP). Results: OP-1 addition obviously decreased cell apoptosis ratio and caspase-3/9 activity, down-regulated gene/protein expression of pro-apoptosis molecules (Bax, caspase-3/cleaved casepase-3 and cleaved PARP), up-regulated gene/protein expression of anti-apoptosis molecule (Bcl-2) in a hyperosmotic culture. Moreover, OP-1 addition significantly increased protein expression of p-Akt and p-mTOR. Further analysis showed that addition of LY294002 and FK-506 partly attenuated these protective effects of OP-1 against NP cell apoptosis and activation of the PI3K/Akt/mTOR pathway in a hyperosmotic culture. Conclusion: OP-1 can attenuate NP cell apoptosis through activating the PI3K/Akt/mTOR pathway in a hyperosmotic culture. The present study sheds a new light on the protective role of OP-1 in regulating disc cell biology and provides some theoretical basis for the application of OP-1 in retarding/regenerating disc degeneration.

scholarly journals Transforming growth factor-β1-regulated Fas/FasL pathway activation suppresses nucleus pulposus cell apoptosis in an inflammatory environment

2020 ◽  
Vol 40 (2) ◽  
Author(s):  
Jingjing Xie ◽  
Bo Li ◽  
Bing Yao ◽  
Pingchao Zhang ◽  
Lixin Wang ◽  
...  
Keyword(s):  
Growth Factor ◽  
Nucleus Pulposus ◽  
Disc Degeneration ◽  
Cell Apoptosis ◽  
Caspase 3 ◽  
Transforming Growth Factor ◽  
Tnf Α ◽  
Gene And Protein Expression ◽  
Tgf Β1

Abstract Background: During disc degeneration, inflammatory cytokine tumor necrosis factor (TNF)-α is correlated with nucleus pulposus (NP) cell apoptosis. Transforming growth factor (TGF)-β1 has the potential to regenerate degenerative disc. Objective: To investigate the protective role of TGF-β1 against TNF-α-mediated NP cell apoptosis and the underlying mechanism. Methods: Rat NP cells were treated with TNF-α (100 ng/ml) for 48 h. TGF-β1 was added into the culture medium to investigate its protective effects against TNF-α-induced NP cell apoptosis. Exogenous FasL was used to investigate the potential role of the Fas/FasL pathway in this process. Flow cytometry assay was used to analyze NP cell apoptosis. Real-time PCR and Western blotting were used to analyze gene and protein expression of apoptosis-related molecules. Results: In TNF-α-treated NP cells, TGF-β1 significantly decreased NP cell apoptosis, declined caspase-3 and -8 activity, and decreased expression of Bax and caspase-3 (cleaved-caspase-3) but increased expression of Bcl-2. However, exogenous FasL partly reversed these effects of TGF-β1 in NP cells treated with TNF-α. Additionally, expression of Fas and FasL in TNF-α-treated NP cells partly decreased by TGF-β1, whereas exogenous FasL increased expression of Fas and FasL in NP cells treated with TGF-β1 and TNF-α. Conclusion: TGF-β1 helps to inhibit TNF-α-induced NP cell apoptosis and the Fas/FasL pathway may be involved in this process. The present study suggests that TGF-β1 may be effective to retard inflammation-mediated disc degeneration.

scholarly journals Resveratrol attenuates mechanical compression-induced nucleus pulposus cell apoptosis through regulating the ERK1/2 signaling pathway in a disc organ culture

2018 ◽  
Vol 38 (2) ◽  
Author(s):  
Zhiwen Zhang ◽  
Feng Wen ◽  
Chengjian He ◽  
Jun Yu
Keyword(s):  
Organ Culture ◽  
Nucleus Pulposus ◽  
Cell Apoptosis ◽  
Caspase 3 ◽  
Dependent Manner ◽  
Protective Effects ◽  
Mechanical Compression ◽  
High Magnitude ◽  
Regulated Expression ◽  
Dose Dependent

Background: Nucleus pulposus (NP) cell apoptosis is a typical feature within the degenerative disc. High magnitude compression significantly promotes NP cell apoptosis. Several studies have indicated that resveratrol has protective effects on disc cell’s normal biology. Objective: The present study aims to investigate whether resveratrol can attenuate mechanical overloading-induced NP cell apoptosis in a disc organ culture. Methods: Isolated porcine discs were cultured in culture chambers of a mechanically active perfusion bioreactor and subjected to a relatively high magnitude compression (1.3 MPa at a frequency of 1.0 Hz for 2 h once per day) for 7 days. Different concentrations (50 and 100 μM) of resveratrol were added into the culture medium to observe the protective effects of resveratrol against NP cell apoptosis under mechanical compression. The noncompressed discs were used as controls. Results: Similar with the previous studies, this high magnitude compression significantly promoted NP cell apoptosis, reflected by the increased number of terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining-positive NP cells and enzyme (caspase-9 and caspase-3) activity, the up-regulated expression of proapoptotic molecules (Bax and caspase-3/cleaved caspase-3), and down-regulated expression of antiapoptotic molecule (Bcl-2). However, resveratrol partly attenuated NP cell apoptosis under this high magnitude compression in a dose-dependent manner. Additionally, though the ERK1/2 pathway was significantly activated in the mechanical compression group, resveratrol partly attenuated activation of the ERK1/2 pathway under mechanical compression in a dose-dependent manner. Conclusion: Resveratrol attenuates mechanical overloading-induced NP cell apoptosis in a dose-dependent manner, and inhibiting activation of the ERK1/2 pathway may be one potential mechanism behind this regulatory process.

scholarly journals GSK-3β Inhibitor SB216763 Resists Oxidative Stress-Induced Apoptosis in Nucleus Pulposus Cell

2021 ◽  
Author(s):  
Kai Zhu ◽  
Song Guo ◽  
Guoyi Han ◽  
Xiancheng Qiang ◽  
Mengmeng Ma ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Membrane Potential ◽  
Nucleus Pulposus ◽  
Mitochondrial Membrane Potential ◽  
Cell Apoptosis ◽  
Mitochondrial Membrane ◽  
Specific Inhibitor ◽  
Nucleus Pulposus Cell ◽  
Gsk 3Β

Abstract Oxidative stress in the intervertebral disc leads to nucleus pulposus (NP) degeneration by inducing cell apoptosis. However, the molecular mechanisms underlying this process remain unclear. Increasing evidence indicates that GSK-3β is related to cell apoptosis induced by oxidative stress. In this study, we explored whether GSK-3β inhibition protects human NP cell against apoptosis under oxidative stress. Immunofluorescence staining was used to show the expression of GSK-3β in human NP cells (NPCs). Flow cytometry, mitochondrial staining and western blot were used to detect apoptosis of treated NPCs, changes of mitochondrial membrane potential and the expression of mitochondrial apoptosis-related proteins using GSK-3β specific inhibitor SB216763. Coprecipitation was used to demonstrate the interaction between GSK-3β and Bcl-2 in an GSK-3β knockdown in vitro model. We delineated the protective effect of GSK-3β specific inhibitor SB216763 on human NP cell apoptosis induced by oxidative stress in vitro. Further, we showed SB216763 exert the protective effect by preservation of the mitochondrial membrane potential and inhibition of caspase 3/7 activity during oxidative injury. The detailed mechanism underlying the antiapoptotic effect of GSK-3β inhibition was also studied by analyzing mitochondrial apoptosis pathway in vitro. We concluded that the GSK-3β inhibitor SB216763 protected mitochondrial membrane potential to delay nucleus pulposus cell apoptosis by inhibiting the interaction between GSK-3β and Bcl-2 and subsequently reducing Cyto-C release and caspase-3 activation. Together, inhibition of GSK-3β using SB216763 in NP may be a favorable therapeutic strategy to slow intervertebral disc degeneration.

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