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
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
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miR-666-3p 通过丝裂原激活蛋白激酶途径介导间充质干细胞来源的外泌体对脑微血管内皮细胞氧葡萄糖剥夺和复氧诱导的细胞损伤的保护作用
DOI：
10.2174/1567202618666210319152534
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发表时间：
2021-01-01
影响因子：
2.1
通讯作者：
Tang, Zhi-xian
中科院分区：
文献类型：
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作者：
Kong, Li-yun;Li, Yan;Tang, Zhi-xian
背景：已有研究报道间充质干细胞(MSC)来源的外切体对原代培养的大鼠脑微血管内皮细胞(BMECs)具有保护作用。目的：探讨MSC来源的外切体对体外培养的大鼠脑微血管内皮细胞(BMEC)的保护作用。进行小RNA深度测序后，进行定量逆转录聚合酶链式反应以验证microRNA(MiRNA)的表达。分别采用细胞计数试剂盒8法、流式细胞术和酶联免疫吸附试验检测RNO-miR-666-3p对OGD/R暴露细胞存活率、细胞凋亡率和炎症反应的影响。此外，我们还利用mRNA深度测序的方法研究了RNO-miR-666-3p在OGD/R细胞基因表达调控中的作用。结果：MSC来源的外切体改变了OGD/R暴露的BMECs的miRNA表达模式。尤其是，RNO-miR-666-3p、RNO-miR-92a-2-5p和ronomiR-219a-2-3p在OGD/R暴露的细胞中的表达水平低于对照组，而MSC衍生的外切体在OGD/R条件下恢复了这些miRNAs的表达水平。过表达RNO-miR-666-3p可提高OGD/R暴露细胞的存活率，减轻细胞凋亡。此外，RNO-miR-666-3p可抑制OGD/R诱导的炎症反应。MRNA深度测序表明，RNO-miR-666-3p与MAPK信号通路密切相关。Western blotting和双荧光素酶分析证实MAPK1是ronomiR-666-3的靶标。结论：MSC来源的外切体恢复了OGD/R暴露的BMEC中RNO-miR-666-3p的表达。此外，这种特异的miRNA通过抑制MAPK信号通路而发挥对OGD/R的保护作用。
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. Naive 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.