Hemeoxygenase-1 Suppresses IL-1β-Induced Apoptosis Through the NF-κB Pathway in Human Degenerative Nucleus Pulposus Cells

Background/Aims: Nucleus pulposus cell (NPC) apoptosis is the main factor in intervertebral disc degeneration (IDD); thus, inhibiting the excessive apoptosis of nucleus pulposus cells may be a potential way to alleviate IDD. The effect of Hemeoxygenase-1 (HO-1) on human NPC apoptosis has never been reported. Our study aimed to investigate the effect and mechanism of HO-1 on apoptosis in human degenerative NPCs. Methods: Nucleus pulposus tissues were collected from patients with lumbar vertebral fracture (LVF) and IDD. The expression of HO-1 and P65 in intervertebral discs was determined using immunohistochemistry and western blot analysis. Apoptosis of human nucleus pulposus cells was quantified by flow cytometric analysis. A recombinant lentiviral vector overexpressing HO-1 and HO-1-siRNA was used to promote or silence the expression of HO-1 in nucleus pulposus cells. The NF-κB inhibitor PDTC was used to inhibit the NF-κB pathway. Results: Our study demonstrated that compared with normal samples, IDD samples showed down-regulation of HO-1 expression and up-regulation of P65 expression. Overexpression of HO-1 inhibited the increase in nucleus pulposus cell apoptosis after IL-1β treatment and simultaneously inhibited the expression of p-P65. Furthermore, after treatment with PDTC, the number of apoptotic cells was significantly decreased with or without overexpression of HO-1. Conclusion: HO-1 might play a significant role in IDD, and HO-1 protected degenerative human NPCs against apoptosis induced by IL-1β through the NF-κB pathway. These findings would aid in the development of novel therapeutic approaches for IDD treatment.

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

10.21203/rs.3.rs-40034/v1 ◽

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).

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Endogenous Bioactive Proteins in Three-Dimensional Cultured Nucleus Pulposus Cells of Herniated Intervertebral Discs: Identification of Potential Target Proteins for Gene Therapeutic Approaches

Global Spine Journal ◽

10.1055/s-0034-1376660 ◽

2014 ◽

Vol 4 (1_suppl) ◽

pp. s-0034-1376660-s-0034-1376660

Author(s):

A. A. Hegewald ◽

J. Fontana ◽

A. Beierfuß ◽

C. Thomé ◽

D. Mern

Keyword(s):

Nucleus Pulposus ◽

Three Dimensional ◽

Intervertebral Discs ◽

Therapeutic Approaches ◽

Nucleus Pulposus Cells ◽

Target Proteins ◽

Potential Target ◽

Bioactive Proteins

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Icariin Prevents IL-1β-Induced Apoptosis in Human Nucleus Pulposus via the PI3K/AKT Pathway

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2017/2198323 ◽

2017 ◽

Vol 2017 ◽

pp. 1-12 ◽

Cited By ~ 7

Author(s):

Xiangyu Deng ◽

Wei Wu ◽

Hang Liang ◽

Donghua Huang ◽

Doudou Jing ◽

...

Keyword(s):

Nucleus Pulposus ◽

Chinese Herb ◽

Specific Inhibitor ◽

Intervertebral Discs ◽

Akt Pathway ◽

Nucleus Pulposus Cells ◽

D 20 ◽

Induced Apoptosis ◽

Epimedium Sagittatum ◽

Lumbar Spondylolisthesis

Purpose. To explore the effect and possible mechanism of icariin, a prenylated flavonol glycoside derived from the Chinese herb Epimedium sagittatum that was applied to IL-1β pretreated human nucleus pulposus (NP) cells. Methods. Human NP cells were isolated from intervertebral discs of patients with scoliosis and lumbar spondylolisthesis. The cells were divided into five groups: A (blank control); B (20 ng/ml IL-1β); C (20 ng/ml IL-1β + 20 μM icariin); D (20 μM icariin + 20 ng/ml IL-1β + 25 μM LY294002); E (20 ng/ml IL-1β + 25 μM LY294002). For each of the five groups, the CCK8, apoptosis rates, ROS rates, and JC-1 rates were determined and an electron micrograph was performed. Different expression levels of apoptosis proteins and proteins in the PI3K/AKT pathway were detected via western blot. Results. We found that the damage effects on human nucleus pulposus cells from 20 ng/ml of IL-1β exposure were attenuated by icariin. When the PI3K/AKT pathway was blocked by LY294002, a specific inhibitor of this pathway, the protective effect of icariin was impaired. In summary, icariin might be a protective traditional Chinese medicine, which prevents inflammation-induced degeneration of intervertebral discs partly through the PI3K/AKT pathway.

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A Combinatorial Relative Mass Value Evaluation of Endogenous Bioactive Proteins in Three-Dimensional Cultured Nucleus Pulposus Cells of Herniated Intervertebral Discs: Identification of Potential Target Proteins for Gene Therapeutic Approaches

PLoS ONE ◽

10.1371/journal.pone.0081467 ◽

2013 ◽

Vol 8 (11) ◽

pp. e81467 ◽

Cited By ~ 14

Author(s):

Demissew S. Mern ◽

Johann Fontana ◽

Anja Beierfuß ◽

Claudius Thomé ◽

Aldemar A. Hegewald

Keyword(s):

Nucleus Pulposus ◽

Three Dimensional ◽

Intervertebral Discs ◽

Relative Mass ◽

Therapeutic Approaches ◽

Nucleus Pulposus Cells ◽

Target Proteins ◽

Value Evaluation ◽

Potential Target ◽

Bioactive Proteins

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Exosomes Derived from Bone Mesenchymal Stem Cells Alleviate Compression-Induced Nucleus Pulposus Cell Apoptosis by Inhibiting Oxidative Stress

Oxidative Medicine and Cellular Longevity ◽

10.1155/2021/2310025 ◽

2021 ◽

Vol 2021 ◽

pp. 1-12

Author(s):

Yiqiang Hu ◽

Ranyang Tao ◽

Linfang Wang ◽

Lang Chen ◽

Ze Lin ◽

...

Keyword(s):

Oxidative Stress ◽

Stem Cells ◽

Mesenchymal Stem Cells ◽

Nucleus Pulposus ◽

Cell Apoptosis ◽

Nucleus Pulposus Cell ◽

Inhibitory Effect ◽

Bone Mesenchymal Stem Cells ◽

Induced Apoptosis ◽

Ivd Degeneration

Oxidative stress is relevant in compression-induced nucleus pulposus (NP) cell apoptosis and intervertebral disc (IVD) degeneration. Exosomes derived from bone mesenchymal stem cells (BMSCs-Exos) are key secretory products of MSCs, with important roles in tissue regeneration. This research is aimed at studying the protective impact of BMSCs-Exos on NP cell apoptosis caused by compression and investigating the underlying mechanisms. Our results indicated that we isolated BMSCs successfully. Exosomes were isolated from the BMSCs and found to alleviate the inhibitory effect that compression has on proliferation and viability in NP cells, decreasing the toxic effects of compression-induced NP cells. AnnexinV/PI double staining and TUNEL assays indicated that the BMSCs-Exos reduced compression-induced apoptosis. In addition, our research found that BMSCs-Exos suppressed compression-mediated NP oxidative stress by detecting the ROS and malondialdehyde level. Furthermore, BMSCs-Exos increased the mitochondrial membrane potential and alleviated compression-induced mitochondrial damage. These results indicate that BMSCs-Exos alleviate compression-mediated NP apoptosis by suppressing oxidative stress, which may provide a promising cell-free therapy for treating IVD degeneration.

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Bone Mesenchymal Stem Cells Promote Extracellular Matrix Remodeling of Degenerated Nucleus Pulposus Cells via the miR-101-3p/EIF4G2 Axis

Frontiers in Bioengineering and Biotechnology ◽

10.3389/fbioe.2021.642502 ◽

2021 ◽

Vol 9 ◽

Author(s):

Zeng Wang ◽

Xiaolin Ding ◽

Feifei Cao ◽

Xishan Zhang ◽

Jingguo Wu

Keyword(s):

Extracellular Matrix ◽

Nucleus Pulposus ◽

Quantitative Polymerase Chain Reaction ◽

Matrix Remodeling ◽

Nucleus Pulposus Cells ◽

Promoting Effect ◽

Bone Mesenchymal Stem Cells ◽

Ecm Remodeling ◽

Lumbar Vertebral Fracture ◽

Related Proteins

The etiology of lumbocrural pain is tightly concerned with intervertebral disk degeneration (IDD). Bone mesenchymal stem cell (BMSC)-based therapy bears potentials for IDD treatment. The properties of microRNA (miRNA)-modified BMSCs may be altered. This study investigated the role and mechanism of BMSCs promoting extracellular matrix (ECM) remodeling of degenerated nucleus pulposus cells (NPCs) via the miR-101-3p/EIF4G2 axis. NPCs were collected from patients with IDD and lumbar vertebral fracture (LVF). The expressions of miR-101-3p and ECM-related proteins, Collagen-I (Col-I) and Collagen-II (Col-II), were detected using the reverse transcription-quantitative polymerase chain reaction. The expressions of Col-I and Col-II, major non-collagenous component Aggrecan, and major catabolic factor Matrix metalloproteinase-13 (MMP-13) were detected using Western blotting. BMSCs were cocultured with degenerated NPCs from patients with IDD. Viability and apoptosis of NPCs were measured using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and flow cytometry. After the degenerated NPCs were transfected with the miR-101-3p inhibitor, the expressions of ECM-related proteins, cell viability, and apoptosis were detected. The targeting relationship between miR-101-3p and EIF4G2 was verified. Functional rescue experiments verified the effects of miR-101-3p and EIF4G2 on ECM remodeling of NPCs. Compared with the NPCs of patients with LVF, the degenerated NPCs of patients with IDD showed downregulated miR-101-3p, Col-II, and Aggrecan expressions and upregulated MMP-13 and Col-I expressions. BMSCs increased the expressions of miR-101-3p, Aggrecan, and Col-II, and decreased the expressions of MMP-13 and Col-I in degenerated NPCs. BMSCs enhanced NPC viability and repressed apoptosis. Downregulation of miR-101-3p suppressed the promoting effect of BMSCs on ECM remodeling. miR-101-3p targeted EIF4G2. Downregulation of EIF4G2 reversed the inhibiting effect of the miR-101-3p inhibitor on ECM remodeling. In conclusion, BMSCs increased the miR-101-3p expression in degenerated NPCs to target EIF4G2, thus promoting the ECM remodeling of NPCs.

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