Brain-enriched microRNA-338 (miR-338) is known to play a central role in brain mitochondrial function, however the role of miR-338 in stroke injury remains unknown. This study investigated the role of miR-338 in injury from transient focal cerebral ischemia in mice, and in cell survival and mitochondrial function after in vitro ischemia in astrocyte and neuronal cultures. Pre-treatment of mice with intracerebroventricular injection of miR-338 antagomir 24 h prior to 1 h of middle cerebral artery occlusion (MCAO) significantly reduced infarct size and improved neurological score at both 24 h and 7d after injury. Levels of the miR-338 target cytochrome-c oxidase subunit 4I1 (COX4I1), which plays an essential role in maintaining brain mitochondrial ATP production, were increased in miR-338 antagomir-treated mice. Mouse primary astrocyte cell cultures subjected to glucose deprivation exhibited increased cell survival when pre-treated with miR-338 inhibitor, and greater cell death with miR-338 mimic. Decreased miR-338 levels were associated with increased ATP production, augmented cytochrome c oxidative (CcO) activity and preservation of COX4I1. In vitro protection with miR-338 inhibitor was blocked by concurrent knockdown of COX4I1 with small interfering RNA. Parallel studies in mouse neuronal N2a cultures resulted in preserved ATP content and CcO activity with miR-338 inhibition, indicating a shared miR-338-dependent response to ischemic stress between brain cell types. These results suggest that miR-338 inhibition and/or COX4I1-targeted therapies may be novel clinical strategies to protect against stroke injury via preservation of mitochondrial function in multiple cell types.

已知大脑中富含的 microRNA-338 (miR-338) 在脑线粒体功能中发挥核心作用，但 miR-338 在中风损伤中的作用仍不清楚。本研究调查了 miR-338 在小鼠短暂性局灶性脑缺血损伤中的作用，以及在星形胶质细胞和神经元培养物中体外缺血后细胞存活和线粒体功能中的作用。在大脑中动脉闭塞 (MCAO) 1 小时前 24 小时，对小鼠脑室内注射 miR-338 antagomir 进行预处理，可显着减少梗死面积，并改善损伤后 24 小时和 7 天的神经系统评分。在 miR-338 antagomir 治疗的小鼠中，miR-338 靶细胞色素 C 氧化酶亚基 4I1 (COX4I1) 的水平有所增加，该亚基在维持脑线粒体 ATP 生成中发挥着重要作用。当用 miR-338 抑制剂预处理时，小鼠原代星形胶质细胞培养物在葡萄糖剥夺条件下表现出细胞存活率增加，而用 miR-338 模拟物预处理时则表现出更大的细胞死亡。 miR-338 水平降低与 ATP 产生增加、细胞色素c氧化 (CcO) 活性增强和 COX4I1 保存相关。 miR-338抑制剂的体外保护作用被小干扰RNA同时敲低COX4I1所阻断。在小鼠神经元 N2a 培养物中进行的平行研究结果表明，miR-338 抑制后 ATP 含量和 CcO 活性得以保留，表明脑细胞类型之间对缺血应激有共同的 miR-338 依赖性反应。这些结果表明，miR-338 抑制和/或 COX4I1 靶向疗法可能是通过保留多种细胞类型中线粒体功能来预防中风损伤的新型临床策略。