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Melatonin alleviates oxidative stress-induced injury to nucleus pulposus-derived mesenchymal stem cells through
Ze-Nan Huang1,2, Ze-Yu Wang3, Xiao-Fei Cheng4
1Department of Orthopedics, Shandong First Medical University & Shandong Academy of Medical Science, Shandong, 200072, China.
Background:
The changes in the microenvironment of degenerative intervertebral discs cause oxidative stress injury and excessive apoptosis of intervertebral disc endogenous stem cells. The purpose of this study was to explore the possible mechanism of the protective effect of melatonin on oxidative stress injury in NPMSCs induced by H2O2.
Methods:
The Cell Counting Kit-8 assay was used to evaluate the cytotoxicity of hydrogen peroxide and the protective effects of melatonin. ROS content was detected by 2'7'-dichlorofluorescin diacetate (DCFH-DA). Mitochondrial membrane potential (MMP) was detected by the JC-1assay. Transferase mediated d-UTP Nick end labeling (TUNEL) and Annexin V/PI double staining were used to determine the apoptosis rate. Additionally, apoptosis-associated proteins and PI3K/Akt signaling pathway-related proteins were evaluated by immunofluorescence, immunoblotting and PCR. ECMs were evaluated by RT‒PCR and immunofluorescence. In vivo, X-ray, Magnetic resonance imaging (MRI) and Histological analyses were used to evaluate the protective effect of melatonin.
Results:
Melatonin had an obvious protective effect on NPMSCs treated with 0-10 μM melatonin for 24 h. In addition, melatonin also had obvious protective effects on mitochondrial dysfunction, decreased membrane potential and cell senescence induced by H2O2. More importantly, melatonin could significantly reduce the apoptosis of nucleus pulposus mesenchymal stem cells induced by H2O2 by regulating the expression of apoptosis-related proteins and decreasing the rate of apoptosis. After treatment with melatonin, the PI3K/Akt pathway was significantly activated in nucleus pulposus mesenchymal stem cells, while the protective effect was significantly weakened after PI3K-IN-1 treatment. In vivo, the results of X-ray, MRI and histological analyses showed that therapy with melatonin could partially reduce the degree of intervertebral disc degeneration.
Conclusion:
Our research demonstrated that melatonin can effectively alleviate the excessive apoptosis and mitochondrial dysfunction of nucleus pulposus mesenchymal stem cells induced by oxidative stress via the PI3K/Akt pathway, which provides a novel idea for the therapy of intervertebral disc degeneration.
The Translational Potential Of This Article:
This study indicates that melatonin can effectively alleviate the excessive apoptosis and mitochondrial dysfunction of NPMSCs through activating the PI3K/Akt pathway. Melatonin might serve as a promising candidate for the prevention and treatment of Intervertebral disc degeneration disease (IVDD) in the future.
Insights
Melatonin protects nucleus pulposus mesenchymal stem cells from oxidative stress and apoptosis by activating the PI3K/Akt pathway. This finding offers a new therapeutic strategy for intervertebral disc degeneration.
Area of Science:
- Biomedical Sciences
- Regenerative Medicine
- Stem Cell Biology
Background:
- Intervertebral disc degeneration (IVDD) involves oxidative stress and apoptosis of nucleus pulposus mesenchymal stem cells (NPMSCs).
- Understanding protective mechanisms against IVDD is crucial for developing effective treatments.
Purpose of the Study:
- To investigate the protective mechanism of melatonin against hydrogen peroxide (H₂O₂)-induced oxidative stress and apoptosis in NPMSCs.
- To explore the role of the PI3K/Akt signaling pathway in melatonin's protective effects.
Main Methods:
- Cell Counting Kit-8 assay for cytotoxicity and protective effects.
- Detection of reactive oxygen species (ROS), mitochondrial membrane potential (MMP), and apoptosis (TUNEL, Annexin V/PI).
- Analysis of apoptosis-related proteins, PI3K/Akt pathway proteins, and extracellular matrix (ECM) components via molecular techniques. In vivo evaluation using X-ray, MRI, and histology.
Main Results:
- Melatonin (0-10 μM) demonstrated significant protective effects against H₂O₂-induced NPMSC damage, including mitochondrial dysfunction and apoptosis.
- Melatonin treatment activated the PI3K/Akt pathway, which was crucial for its protective effects, as evidenced by PI3K-IN-1 inhibition.
- In vivo studies showed melatonin partially mitigated intervertebral disc degeneration.
Conclusions:
- Melatonin effectively alleviates oxidative stress-induced apoptosis and mitochondrial dysfunction in NPMSCs by activating the PI3K/Akt pathway.
- Melatonin presents a promising therapeutic candidate for preventing and treating intervertebral disc degeneration.
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