Ischemic stroke is the major form of stroke and is accentuated by multiple comorbidities. It has been previously shown that different microRNAs (miRNAs) regulate separate aspects of ischemic stroke. Differential miRNA expression analysis in cerebrospinal fluid of stroke patients had revealed upregulation of miR‐124‐3p, miR‐9‐3p, miR‐9‐5p, and miR‐128‐3p. However, whether the overexpression is correlative or causative was not known. Here, using an in vitro oxygen–glucose deprivation/reoxygenation (OGD/R) neuronal cell model, we saw OGD/R‐induced injury was associated with significant upregulation of the aforementioned four miRNAs. Target gene prediction using in situ algorithms and gene set enrichment analysis revealed significant enrichment of FOXO and Relaxin signaling pathways and regulatory processes associated with endothelial cell migration, which are all known to associate with apoptotic pathways. In situ protein–protein interaction network analysis confirmed the findings of gene set enrichment analysis. TUNEL analysis showed that OGD/R‐induced injury resulted in significant apoptosis, which was significantly inhibited in neuronal cells pretransfected with inhibitors of either miR‐9‐5p or miR‐128‐3p. Further testing in an in vivo middle cerebral artery occlusion (MCAO) mouse model of ischemic stroke showed that inhibiting miR‐9‐5p or miR‐128‐3p significantly decreases MCAO‐induced infraction volume and inhibited apoptotic response as revealed by decreased cleaved Caspase‐3 protein expression in immunohistochemical analysis. Combined inhibition of miR‐9‐5p and miR‐128‐3p resulted in a synergistic decrease in cell death and infraction volume in vitro and in vivo, respectively. Cumulatively, our results provide critical knowledge about the mechanism by which elevated miR‐9‐5p and miR‐128‐3p causes brain damage in ischemic stroke and provides evidence of them being attractive therapeutic targets.

中文翻译：

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抑制microRNA-9-5p和microRNA-128-3p可在体外和体内抑制缺血性卒中相关细胞死亡

缺血性中风是中风的主要形式，并因多种合并症而加重。先前已经表明，不同的微 RNA (miRNA) 调节缺血性中风的不同方面。中风患者脑脊液中的差异 miRNA 表达分析显示 miR-124-3p、miR-9-3p、miR-9-5p 和 miR-128-3p 上调。然而，过表达是相关的还是因果关系尚不清楚。在这里，使用体外氧-葡萄糖剥夺/复氧 (OGD/R) 神经元细胞模型，我们发现 OGD/R 诱导的损伤与上述四种 miRNA 的显着上调有关。使用原位算法和基因集富集分析的靶基因预测揭示了与内皮细胞迁移相关的 FOXO 和松弛素信号通路和调控过程的显着富集，众所周知，这些都与凋亡途径有关。原位蛋白质-蛋白质相互作用网络分析证实了基因集富集分析的结果。TUNEL 分析表明，OGD/R 诱导的损伤导致显着的细胞凋亡，这在用 miR-9-5p 或 miR-128-3p 抑制剂预转染的神经元细胞中受到显着抑制。在缺血性卒中的体内大脑中动脉闭塞 (MCAO) 小鼠模型中的进一步测试表明，抑制 miR-9-5p 或 miR-128-3p 可显着降低 MCAO 诱导的梗死体积并抑制细胞凋亡反应，如裂解的 Caspase- 3 免疫组化分析中的蛋白质表达。miR-9-5p 和 miR-128-3p 的联合抑制分别导致体外和体内细胞死亡和损伤体积的协同减少。