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miR-666-3p Mediates the Protective Effects of Mesenchymal Stem Cell-derived Exosomes Against Oxygen-glucose Deprivation and Reoxygenation- induced Cell Injury in Brain Microvascular Endothelial Cells via Mitogen-activated Protein Kinase Pathway

[ Vol. 18 , Issue. 1 ]

Author(s):

Li-yun Kong, Yan Li, Ding-yu Rao, Bing Wu, Cheng-peng Sang, Ping Lai, Jun-song Ye, Zu-xiong Zhang, Zhi-ming Du, Jun-jian Yu, Liang Gu, Fa-chun Xie, Zi-you Liu* and Zhi-xian Tang*   Pages 20 - 77 ( 58 )

Abstract:


Background: Previous studies have reported that mesenchymal stem cell (MSC)- derived exosomes can protect primary rat brain microvascular endothelial cells (BMECs) against oxygen-glucose deprivation and reoxygenation (OGD/R)-induced injury.

Objective: The aim was to identify the key factors mediating the protective effects of MSC-derived exosomes.

Methods: Primary rat BMECs were either pretreated or not pretreated with MSC-derived exosomes before exposure to OGD/R. Naïve cells were used as a control. After performing small RNA deep sequencing, quantitative reverse transcription polymerase chain reaction was performed to validate microRNA (miRNA) expression. The effects of rno-miR-666-3p on cell viability, apoptosis, and inflammation in OGD/R-exposed cells were assessed by performing the Cell Counting Kit 8 assay, flow cytometry, and enzyme-linked immunosorbent assay, respectively. Moreover, the role of rno-miR-666-3p in regulating gene expression in OGD/R-exposed cells was studied using mRNA deep sequencing. Lastly, to evaluate whether mitogen-activated protein kinase 1 (MAPK1) was the target of rno-miR-666-3p, western blotting and the dual-luciferase assay were performed.

Results: MSC-derived exosomes altered the miRNA expression patterns in OGD/R-exposed BMECs. In particular, the expression levels of rno-miR-666-3p, rno-miR-92a-2-5p, and rnomiR- 219a-2-3p decreased in OGD/R-exposed cells compared with those in the control; however, MSC-derived exosomes restored the expression levels of these miRNAs under OGD/R conditions. rno-miR-666-3p overexpression enhanced cell viability and alleviated the apoptosis of OGD/R-exposed cells. Moreover, rno-miR-666-3p suppressed OGD/R-induced inflammation. mRNA deep sequencing revealed that rno-miR-666-3p is closely associated with the MAPK signaling pathway. Western blotting and the dual-luciferase assay confirmed that MAPK1 is the target of rnomiR- 666-3p.

Conclusion: MSC-derived exosomes restore rno-miR-666-3p expression in OGD/R-exposed BMECs. Moreover, this specific miRNA exerts protective effects against OGD/R by suppressing the MAPK signaling pathway.

Keywords:

Oxygen–glucose deprivation, brain, endothelial cells, mesenchymal stem cells, exosomes, microRNAs.

Affiliation:

Department of Operation Room, Heart Center, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, Department of Clinical Pharmacy, The First Affiliated Hospital of Gannan Medical University, Guanzhou, 341000, Department of Cardiothoracic Surgery, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, Department of Anatomy, Gannan Medical University, Guanzhou, 341000, Department of Cardiothoracic Surgery, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, Department of Cardiology, Heart Center, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, Department of Clinical Research Center, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, Department of Cardiothoracic Surgery, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, Department of Cardiothoracic Surgery, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, Department of Cardiothoracic Surgery, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, Department of Cardiothoracic Surgery, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, Department of Cardiothoracic Surgery, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, Department of Cardiothoracic Surgery, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, Department of Cardiothoracic Surgery, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000



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