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

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.

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Mechano growth factor attenuates mechanical overload-induced nucleus pulposus cell apoptosis through inhibiting the p38 MAPK pathway

Bioscience Reports ◽

10.1042/bsr20182462 ◽

2019 ◽

Vol 39 (3) ◽

Cited By ~ 4

Author(s):

Qing Xu ◽

Haolin Fang ◽

Liang Zhao ◽

Cunxin Zhang ◽

Luo Zhang ◽

...

Keyword(s):

Growth Factor ◽

Protein Expression ◽

Nucleus Pulposus ◽

P38 Mapk ◽

Disc Degeneration ◽

Cell Apoptosis ◽

Caspase 3 ◽

Mapk Pathway ◽

P38 Mapk Pathway ◽

Mechanical Overload

Abstract Mechanical overload is a risk factor of disc degeneration. It can induce disc degeneration through mediating cell apoptosis. Mechano growth factor (MGF) has been reported to inhibit mechanical overload-induced apoptosis of chondrocytes. The present study is aimed to investigate whether MGF can attenuate mechanical overload-induced nucleus pulposus (NP) cell apoptosis and the possible signaling transduction pathway. Rat NP cells were cultured and subjected to mechanical overload for 7 days. The control NP cells did not experience mechanical load. The exogenous MGF peptide was added into the culture medium to investigate its protective effects. NP cell apoptosis ratio, caspase-3 activity, gene expression of Bcl-2, Bax and caspase-3, protein expression of cleaved caspase-3, cleaved PARP, Bax and Bcl-2 were analyzed to evaluate NP cell apoptosis. In addition, activity of the p38 MAPK pathway was also detected. Compared with the control NP cells, mechanical overload significantly increased NP cell apoptosis and caspase-3 activity, up-regulated gene/protein expression of pro-apoptosis molecules (i.e. Bax, caspase-3, cleaved caspase-3 and cleaved PARP) whereas down-regulated gene/protein expression of anti-apoptosis molecule (i.e. Bcl-2). However, exogenous MGF partly reversed these effects of mechanical overload on NP cell apoptosis. Further results showed that activity of the p38 MAPK pathway of NP cells cultured under mechanical overload was decreased by addition of MGF peptide. In conclusion, MGF is able to attenuate mechanical overload-induced NP cell apoptosis, and the p38 MAPK signaling pathway may be involved in this process. The present study provides that MGF supplementation may be a promising strategy to retard mechanical overload-induced disc degeneration.

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Role of p38–MAPK pathway in the effects of high-magnitude compression on nucleus pulposus cell senescence in a disc perfusion culture

Bioscience Reports ◽

10.1042/bsr20170718 ◽

2017 ◽

Vol 37 (5) ◽

Cited By ~ 14

Author(s):

Lianglong Pang ◽

Pei Li ◽

Ruijie Zhang ◽

Yuan Xu ◽

Lei Song ◽

...

Keyword(s):

Nucleus Pulposus ◽

P38 Mapk ◽

Ex Vivo ◽

Mapk Pathway ◽

Cell Senescence ◽

Bioreactor Culture ◽

Pathological Feature ◽

High Magnitude ◽

P38 Mapk Pathway

Nucleus pulposus (NP) cell senescence is a typical pathological feature within the degenerative intervertebral disc. As a potential inducing and aggregating factor of disc degeneration, mechanical overloading affects disc biology in multiple ways. The present study was to investigate the NP cell senescence-associated phenotype under intermittent high compression in an ex vivo disc bioreactor culture, and the role of the p38–MAPK pathway in this regulatory process. Porcine discs were cultured in culture chambers of a self-developed mechanically active bioreactor and subjected to different magnitudes of dynamic compression (low-magnitude and high-magnitude: 0.1 and 1.3 MPa at a frequency of 1.0 Hz for 2 h per day respectively) for 7 days. Non-compressed discs were used as controls. The inhibitor SB203580 was used to study the role of the p38–MAPK pathway in this process. Results showed that intermittent high-magnitude compression clearly induced senescence-associated changes in NP cells, such as increasing β-galactosidase-positive NP cells, decreasing PCNA-positive NP cells, promoting the formation of senescence-associated heterochromatic foci (SAHF), up-regulating the expression of senescence markers (p16 and p53), and attenuating matrix production. However, inhibition of the p38–MAPK pathway partly attenuated the effects of intermittent high-magnitude (1.3 MPa) compression on those described NP cell senescence-associated parameters. In conclusion, intermittent high-magnitude compression can induce NP cell senescence-associated changes in an ex vivo disc bioreactor culture, and the p38–MAPK pathway is involved in this process.

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Acidic pH promotes nucleus pulposus cell senescence through activating the p38 MAPK pathway

Bioscience Reports ◽

10.1042/bsr20181451 ◽

2018 ◽

Vol 38 (6) ◽

Cited By ~ 6

Author(s):

Jiabin Fu ◽

Wei Yu ◽

Dianming Jiang

Keyword(s):

Nucleus Pulposus ◽

P38 Mapk ◽

Mapk Pathway ◽

Cell Senescence ◽

Phase Fraction ◽

Control Group ◽

Acidic Ph ◽

Degenerative Disc ◽

P38 Mapk Pathway ◽

Regulated Expression

Background: Nucleus pulposus (NP) cell senescence is an important cellular feature within the degenerative disc. It is known that a very acidic niche exists in the degenerative disc, which participates in regulating disc cell viability and matrix metabolism. Objective: The present study was aimed to investigate the role and potential signaling transduction pathway of an acidic pH in regulating NP cell senescence. Methods: Rat NP cells were cultured in an acidic pH of 7.2 close to that in a healthy disc (Control group) or in an acidic pH of 6.2 close to that in a severe degenerative disc (Experiment group) for 10 days. Additionally, the experimental NP cells were incubated along with the inhibitor SB203580 to analyze the role of p38 MAPK pathway in this process. Results: Compared with the control NP cells, experimental NP cells showed a suppressed cell proliferation potency, an increased G0/G1 phase fraction whereas a decreased S-phase fraction and a declined telomerase activity, an up-regulated expression of senescence-related molecules (p16 and p53), and a down-regulated expression of matrix-related moleucles (aggrecan and collagen II). Further analysis showed that inhibition of the p38 MAPK pathway partly reversed effects of acidic pH of 6.2 on the experimental NP cells. Conclusion: The very acidic niche identified in a severe degenerative disc promotes NP cell senescence through regulating the p38 MAPK pathway. The present study provides a new mechanism that drives NP cell senescence during disc degeneration.

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Aquaporin-3 Attenuates Oxidative Stress-Induced Nucleus Pulposus Cell Apoptosis Through Regulating the P38 MAPK Pathway

Cellular Physiology and Biochemistry ◽

10.1159/000494788 ◽

2018 ◽

Vol 50 (5) ◽

pp. 1687-1697 ◽

Cited By ~ 10

Author(s):

Yichun Xu ◽

Hui Yao ◽

Qiyou Wang ◽

Wenbin Xu ◽

Kaihua Liu ◽

...

Keyword(s):

Gene Expression ◽

Oxidative Damage ◽

P38 Mapk ◽

Cell Apoptosis ◽

Caspase 3 ◽

Mapk Pathway ◽

Aquaporin 3 ◽

P38 Mapk Pathway ◽

Intracellular Ros ◽

Cellular Apoptosis

Background/Aims: Previous studies have shown that oxidative damage is a main contributor to disc nucleus pulposus (NP) cell apoptosis. Aquaporin-3 (AQP-3) facilitates reactive oxygen species (ROS) scavenging and thus alleviates oxidative injury in other cells. This study aims to investigate the role and mechanism of AQP-3 in regulating NP cell apoptosis under oxidative damage. Methods: Rat NP cells were treated with H2O2 for 48 hours, while control NP cells were free of H2O2. Recombinant AQP-3 lentiviral vectors were used to investigate the effect of enhanced AQP-3 expression levels in NP cells. NP cell apoptosis was assessed by flow cytometry, caspase-3 activity, gene expression of apoptosis-related molecules (Bax, Bcl-2 and caspase-3), and protein expression of cellular apoptosis markers (cleaved PARP and cleaved caspase-3). Additionally, intracellular ROS content and activity of the p38 MAPK pathway were evaluated. Results: Compared with the control NP cells, oxidative damage in the treatment cells significantly increased cell apoptosis ratios and caspase-3 activity, upregulated gene expression of Bax and caspase-3, downregulated gene expression of Bcl-2, and increased protein expression of cleaved PARP and cleaved caspase-3, as well as increased intracellular ROS content and activity of the p38 MAPK pathway. However, AQP-3 overexpression partly alleviated cell apoptosis, decreased intracellular ROS content, and inhibited the p38 MAPK pathway in NP cells under oxidative damage. Conclusion: Oxidative damage can significantly downregulate AQP-3 expression. Enhancing AQP-3 expression in NP cells partly attenuates cellular apoptosis through regulating the p38 MAPK pathway under oxidative damage.

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High glucose promotes annulus fibrosus cell apoptosis through activating the JNK and p38 MAPK pathways

Bioscience Reports ◽

10.1042/bsr20190853 ◽

2019 ◽

Vol 39 (7) ◽

Cited By ~ 6

Author(s):

Lizhen Shan ◽

Di Yang ◽

Danjie Zhu ◽

Fabo Feng ◽

Xiaolin Li

Keyword(s):

P38 Mapk ◽

High Glucose ◽

Cell Apoptosis ◽

Annulus Fibrosus ◽

Caspase 3 ◽

Mapk Pathway ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Jnk Pathway ◽

Regulated Expression

Abstract Diabetes mellitus (DM) is an important risk factor of intervertebral disc degeneration. A high glucose niche-mediated disc cell apoptosis is an implicate causative factor for the spine degenerative diseases related with DM. However, the effects of a high glucose niche on disc annulus fibrosus (AF) cell apoptosis and the potential signaling transduction pathway is unclear. The present study is to investigate the effects of high glucose on disc AF cell apoptosis and the role of two potential signaling pathways in this process. Rat AF cells were cultured in baseline medium or medium with different concentrations (0.1 and 0.2 M) of glucose for 3 days. Flow cytometry was used to assess the degree of apoptosis. Activity of caspase 3/9 was evaluated by chemical kit. Expression of pro-apoptotic and anti-apoptotic molecules was analyzed by real-time polymerase chain reaction and Western blot. In addition, activity of the C-Jun NH2-terminal kinases (JNK) pathway and p38 mitogen-activated protein kinase (MAPK) pathway was evaluated by Western blot. Compared with the control group, high glucose culture increased cell apoptosis ratio and caspase-3/9 activity, up-regulated expression of bax, caspase-3, cleaved caspase-3 and cleaved PARP, and down-regulated expression of bcl-2 in a glucose concentration-dependent manner. Additionally, high glucose culture increased expression of the p-JNK and p-p38 MAPK in a concentration-dependent manner. Further results showed that inhibition of the JNK or p38 MAPK pathway attenuated the effects of high glucose on AF cell apoptosis. Together, high glucose promoted disc AF cell apoptosis through regulating the JNK pathway and p38 MAPK pathway in a glucose concentration-dependent manner.

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Inflammatory cytokines induce caveolin-1/β-catenin signalling in rat nucleus pulposus cell apoptosis through the p38 MAPK pathway

Cell Proliferation ◽

10.1111/cpr.12254 ◽

2016 ◽

Vol 49 (3) ◽

pp. 362-372 ◽

Cited By ~ 40

Author(s):

Jianxi Wang ◽

Huajiang Chen ◽

Peng Cao ◽

Xiaodong Wu ◽

Fazhi Zang ◽

...

Keyword(s):

Nucleus Pulposus ◽

P38 Mapk ◽

Inflammatory Cytokines ◽

Cell Apoptosis ◽

Mapk Pathway ◽

Nucleus Pulposus Cell ◽

Caveolin 1 ◽

P38 Mapk Pathway

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

Bioscience Reports ◽

10.1042/bsr20171703 ◽

2018 ◽

Vol 38 (2) ◽

Cited By ~ 8

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.

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Qingxin Kaiqiao Fang Inhibits Aβ25-35-Induced Apoptosis in Primary Cultured Rat Hippocampal Neuronal Cells via the p38 MAPK Pathway: An Experimental Validation and Network Pharmacology Study

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2020/9058135 ◽

2020 ◽

Vol 2020 ◽

pp. 1-14

Author(s):

Tian-Qi Wang ◽

Xiao-Xiao Lai ◽

Lu-Ting Xu ◽

Yan Shen ◽

Jian-Wei Lin ◽

...

Keyword(s):

Cell Viability ◽

P38 Mapk ◽

Caspase 3 ◽

Mapk Pathway ◽

Neuronal Cells ◽

Neuronal Morphology ◽

Cell Counting ◽

Network Pharmacology ◽

Expression Levels ◽

P38 Mapk Pathway

Qingxin kaiqiao fang (QKF), a traditional Chinese medicine compound, has been applied to treat Alzheimer’s disease (AD) for many years and has exhibited remarkable effects. However, the underlying mechanism is still not explicit. The current study aims to investigate whether QKF exerts an antiapoptotic role through the p38 MAPK pathway in the course of AD. Network pharmacology analysis was applied to study the effective components, possible therapeutic targets, and AD-related pathway of QKF. Further, the AD cell model was established using amyloid-beta (Aβ)25-35 peptide and primary hippocampal neuronal cells extracted from newborn Sprague-Dawley rats. Microtubule-associated protein-2 (MAP-2) imaging was used to detect the morphology of hippocampal neurons. Western blot (WB) analysis was applied to detect the protein expression levels of p38 MAPK, p-p38 MAPK, Bcl-2, Bax, caspase-3, and cleaved caspase-3. Cell viability and apoptosis were determined using cell counting kit-8 (CCK-8) and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assays, respectively. SB203580 and U46619 were used to detect changes in cell morphology, cell viability, and apoptosis upon inhibiting or activating p38 MAPK. Our present work showed that QKF protects hippocampal neuronal morphology, enhances cell viability, and reduces the number of TUNEL-positive cells. In addition, our results showed that QKF increased the expression levels of antiapoptotic proteins and decreased the expression of proapoptotic proteins. QKF at 25 mg·mL−1 best inhibited neuronal apoptosis among the three doses of QKF by suppressing p38 MAPK activity. Collectively, QKF plays an antiapoptotic role via the p38 MAPK pathway.

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Osteogenic protein-1 alleviates high glucose microenvironment-caused degenerative changes in nucleus pulposus cells

Bioscience Reports ◽

10.1042/bsr20190170 ◽

2019 ◽

Vol 39 (4) ◽

Cited By ~ 3

Author(s):

Ziming Liu ◽

Zhiwen Zhang ◽

Ali Zhang ◽

Fan Zhang ◽

Wennan Du ◽

...

Keyword(s):

High Glucose ◽

Cell Apoptosis ◽

Culture Medium ◽

Caspase 3 ◽

Degenerative Changes ◽

Protective Effects ◽

Regulated Expression ◽

Osteogenic Protein ◽

Collagen Ii ◽

Apoptosis Ratio

Abstract Increasing evidence has indicated a close relationship between diabetes mellitus (DM) and disc degeneration. As a potential therapeutic growth factor, osteogenic protein-1 (OP-1) has lots of protective effects on the healthy disc cell’s biology. The present study was aimed to investigate the effects of OP-1 on degenerative changes of nucleus pulposus (NP) cells in a high glucose culture. Rat NP cells were cultured in the baseline medium or the high glucose (0.2 M) culture medium. OP-1 was added into the high glucose culture medium to investigate whether its has some protective effects against degenerative changes of NP cells in the high glucose culture. NP cell apoptosis ratio, caspase-3/9 activity, expression of apoptosis-related molecules (Bcl-2, Bax, and caspase-3), matrix macromolecules (aggrecan and collagen II), and matrix remodeling enzymes (MMP-3, MMP-13, and ADAMTS-4), and immuno-staining of NP matrix proteins (aggrecan and collagen II) were evaluated. Compared with the baseline culture, high glucose culture significantly increased NP cell apoptosis ratio, caspase-3/9 activity, up-regulated expression of Bax, caspase-3, MMP-3, MMP-13 and ADAMTS-4, down-regulated expression of Bcl-2, aggrecan and collagen II, and decreased staining intensity of aggrecan and collagen II. However, the results of these parameters were partly reversed by the addition of OP-1 in the high glucose culture. OP-1 can alleviate high glucose microenvironment-induced degenerative changes of NP cells. The present study provides that OP-1 may be promising in retarding disc degeneration in DM patients.

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

Bioscience Reports ◽

10.1042/bsr20190043 ◽

2019 ◽

Vol 39 (3) ◽

Cited By ~ 9

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.

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