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miR-9a-5p Protects Ischemic Stroke by Regulating Oxidative Stress and Mitochondrial Autophagy
Chunli Ma1, Qing Gao2, Li Zhang3
1Department of Neurology, The Second Affiliated Hospital of Mudanjiang Medical University, Mudanjiang, 157010 Heilongjiang, China.
Purpose:
Present research is aimed at exploring the effect of miR-9a-5p on mitochondrial autophagy and alleviating cellular oxidative stress injury in ischemic stroke.
Methods:
SH-SY5Y cells were cultured with oxygen-glucose deprivation/reoxygenation (OGD/R) to simulate ischemia/reperfusion. The cells were treated in an anaerobic incubator (95% N2, 5% CO2) for 2 h and then reoxygenated in the normoxic condition for 24 h with 2 ml of normal medium. Cells were transfected with miR-9a-5p mimic/inhibitor or negative control. The RT-qPCR assay was utilized to measure the mRNA expression. Western blot was utilized to evaluate the protein expression. The CCK-8 assay was conducted to detect cell viability. Flow cytometry was applied to examine apoptosis and the cell cycle. The ELISA assay was applied to measure the contents of SOD and MDA in mitochondria. Autophagosomes were observed via electron microscopy.
Results:
By comparison with the control group, the miR-9a-5p expression in the OGD/R group obviously declined. Mitochondrial crista breaks, vacuole-like changes, and increased autophagosome formation were observed in the OGD/R group. OGD/R injury enhanced oxidative stress damage and mitophagy. When transfected with the miR-9a-5p mimic, mitophagosome production of SH-SY5Y cells decreased and oxidative stress injury was inhibited. However, the miR-9a-5p inhibitor obviously increased mitophagosome production and enhanced oxidative stress injury.
Conclusion:
miR-9a-5p protects against ischemic stroke by inhibiting OGD/R-induced mitochondrial autophagy and alleviating cellular oxidative stress injury.
Insights
MicroRNA-9a-5p (miR-9a-5p) plays a protective role in ischemic stroke by reducing mitochondrial autophagy and oxidative stress. Upregulating miR-9a-5p alleviates cell damage, while inhibiting it worsens the injury.
Area of Science:
- Neuroscience
- Molecular Biology
- Cell Biology
Background:
- Ischemic stroke causes significant cellular oxidative stress and mitochondrial damage.
- Mitochondrial autophagy, or mitophagy, is implicated in the progression of ischemic stroke injury.
- MicroRNAs (miRNAs) are key regulators of cellular processes, including stress responses and autophagy.
Purpose of the Study:
- To investigate the role of miR-9a-5p in regulating mitochondrial autophagy.
- To determine the effect of miR-9a-5p on oxidative stress injury in an in vitro model of ischemic stroke.
- To explore miR-9a-5p as a potential therapeutic target for ischemic stroke.
Main Methods:
- Established an in vitro model of ischemic stroke using oxygen-glucose deprivation/reoxygenation (OGD/R) in SH-SY5Y cells.
- Utilized RT-qPCR and Western blot to assess miR-9a-5p expression and protein levels.
- Employed CCK-8, flow cytometry, ELISA, and electron microscopy to evaluate cell viability, apoptosis, oxidative stress markers (SOD, MDA), and autophagosome formation.
Main Results:
- OGD/R significantly decreased miR-9a-5p expression and induced mitochondrial damage, increased mitophagy, and enhanced oxidative stress.
- Transfection with a miR-9a-5p mimic reduced mitophagosome production and alleviated OGD/R-induced oxidative stress injury.
- Conversely, inhibition of miR-9a-5p exacerbated mitophagosome production and oxidative stress.
Conclusions:
- miR-9a-5p exhibits a protective effect against ischemic stroke.
- The mechanism involves the inhibition of OGD/R-induced mitochondrial autophagy and the alleviation of cellular oxidative stress.
- miR-9a-5p represents a promising therapeutic target for mitigating ischemic stroke damage.
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