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Isolating, Sequencing and Analyzing Extracellular MicroRNAs from Human Mesenchymal Stem Cells
Published on: March 8, 2019
Exosomes Released from Bone-Marrow Stem Cells Ameliorate Hippocampal Neuronal Injury Through transferring miR-455-3p
1Department of Anesthesiology, the Third People's Hospital of Hubei Province, Wuhan 430050, Hubei, China.
Background:
The neuroprotective roles of mesenchymal stem cells (MSCs) in brain injury are elicited at least partially through the secretion exosomes containing microRNAs (miRNAs). We herein investigate the protective function of bone marrow MSCs (BMSCs)-derived exosomes harboring miR-455-3p against hippocampal neuronal injury in mouse and N2a cell damage model.
Methods:
First, BMSC surface markers were detected by flow cytometry, followed by extraction of BMSCs-derived exosomes (BMSCs-Exos). A mouse model of neuronal injury was induced by middle cerebral artery occlusion/reperfusion (MCAO/R), and N2a cells were exposed to oxygen-glucose deprivation/reoxygenation (OGD/R) for in vitro experiments. BMSCs-Exos were administrated in mice and N2a cells. We subsequently determined viability- and apoptosis-features using EdU staining, CCK-8, flow cytometry and Caspase-3 kits. Subsequently, we used RT-qPCR to assess miR-455-3p expression in brain tissues as well as N2a cells, and bioinformatic tools to predict the targeting mRNA of miR-455-3p, which was validated by dual-luciferase assays.
Results:
BMSCs-Exos improved hippocampal neuronal injury in MCAO/R-treated mice and OGD/R-induced injury to N2a cells. BMSCs-Exos upregulated miR-455-3p expression in brain tissues of mice and OGD/R-treated N2a cells. miR-455-3p targeted and conversely regulated PDCD7 expression. The protective effect of BMSCs-Exos on OGD/R-treated N2a cells was markedly mitigated following miR-455-3p downregulation. Moreover, overexpression of miR-455-3p contributed to increased N2a cell activity and decreased apoptosis, while the rescue experiment results were opposite.
Conclusion:
MSCs-derived exosomal miR-455-3p targeted PDCD7 to alleviate hippocampal neuronal injury in MCAO/R-treated mice and injury of OGD/R-treated N2a cells.
Insights
Bone marrow mesenchymal stem cells (BMSCs)-derived exosomes protect against brain injury by delivering miR-455-3p. This microRNA targets PDCD7, reducing neuronal damage in models of stroke and cellular injury.
Area of Science:
- Neuroscience
- Stem Cell Biology
- Molecular Biology
Background:
- Mesenchymal stem cells (MSCs) exert neuroprotection via exosome-secreted microRNAs (miRNAs).
- Bone marrow MSCs (BMSCs)-derived exosomes carrying miR-455-3p are investigated for their protective effects.
- Focus on hippocampal neuronal injury models in mice and N2a cells.
Purpose of the Study:
- To investigate the neuroprotective role of BMSCs-derived exosomes harboring miR-455-3p.
- To elucidate the mechanism by which miR-455-3p in exosomes affects neuronal injury.
- To evaluate therapeutic potential in in vivo and in vitro models.
Main Methods:
- BMSC surface markers confirmed by flow cytometry; exosome extraction (BMSCs-Exos).
- In vivo: Middle cerebral artery occlusion/reperfusion (MCAO/R) mouse model.
- In vitro: Oxygen-glucose deprivation/reoxygenation (OGD/R) in N2a cells; BMSCs-Exos administration.
- Assessed cell viability, apoptosis (EdU, CCK-8, flow cytometry, Caspase-3), miR-455-3p expression (RT-qPCR), and target gene validation (dual-luciferase).
Main Results:
- BMSCs-Exos treatment improved neuronal survival and reduced apoptosis in both MCAO/R mice and OGD/R N2a cells.
- Exosomal miR-455-3p was upregulated in brain tissues and N2a cells post-treatment.
- miR-455-3p directly targets and downregulates PDCD7 expression.
- Downregulating miR-455-3p diminished the protective effects of BMSCs-Exos; miR-455-3p overexpression increased cell viability and decreased apoptosis.
Conclusions:
- MSCs-derived exosomal miR-455-3p plays a crucial role in neuroprotection.
- The mechanism involves targeting PDCD7 to alleviate hippocampal neuronal injury.
- BMSCs-Exos represent a promising therapeutic strategy for brain injury.
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