Department of Operation Room, Heart Center, The First Affiliated Hospital of Gannan Medical University, Ganzhou, 341000, China.
Department of Clinical Pharmacy, The First Affiliated Hospital of Gannan Medical University, Guanzhou, 341000, China.
Curr Neurovasc Res. 2021;18(1):20-77. doi: 10.2174/1567202618666210319152534.
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.
The aim was to identify the key factors mediating the protective effects of MSC-derived exosomes.
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.
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.
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.
先前的研究报告称，间充质干细胞（MSC）衍生的外泌体可以保护原代大鼠脑微血管内皮细胞（BMEC）免受氧葡萄糖剥夺和再氧合（OGD/R）诱导的损伤。
旨在确定介导 MSC 衍生的外泌体保护作用的关键因素。
在暴露于 OGD/R 之前，用 MSC 衍生的外泌体预处理或不预处理原代大鼠 BMEC。幼稚细胞用作对照。进行小 RNA 深度测序后，通过定量逆转录聚合酶链反应验证 microRNA（miRNA）表达。通过细胞计数试剂盒 8 测定、流式细胞术和酶联免疫吸附测定分别评估 rno-miR-666-3p 在 OGD/R 暴露细胞中的细胞活力、凋亡和炎症的影响。此外，使用 mRNA 深度测序研究 rno-miR-666-3p 在 OGD/R 暴露细胞中调节基因表达的作用。最后，通过 Western blot 和双荧光素酶测定评估 MAPK1 是否是 rno-miR-666-3p 的靶标。
MSC 衍生的外泌体改变了 OGD/R 暴露的 BMEC 中的 miRNA 表达模式。特别是，与对照组相比，rno-miR-666-3p、rno-miR-92a-2-5p 和 rnomiR-219a-2-3p 的表达水平在 OGD/R 暴露的细胞中降低；然而，MSC 衍生的外泌体在 OGD/R 条件下恢复了这些 miRNA 的表达水平。rno-miR-666-3p 过表达增强了细胞活力并减轻了 OGD/R 暴露细胞的凋亡。此外，rno-miR-666-3p 抑制了 OGD/R 诱导的炎症。mRNA 深度测序显示 rno-miR-666-3p 与 MAPK 信号通路密切相关。Western blot 和双荧光素酶测定证实 MAPK1 是 rnomiR-666-3p 的靶标。
MSC 衍生的外泌体恢复了 OGD/R 暴露的 BMEC 中 rno-miR-666-3p 的表达。此外，这种特定的 miRNA 通过抑制 MAPK 信号通路对 OGD/R 发挥保护作用。
