Inhibition of microRNA-200c preserves astrocyte sirtuin-1 and mitofusin-2, and protects against hippocampal neurodegeneration following global cerebral ischemia in mice.

Memory impairment remains a leading disability in survivors of global cerebral ischemia, occurring secondary to delayed neurodegeneration of hippocampal cornu ammonis-1 (CA1) neurons. MicroRNA-200c (miR-200c) is induced following ischemic stress and we have previously demonstrated that pre-treatment with anti-miR-200c is protective against embolic stroke in mice. In the present study we assessed the role of miR-200c on CA1 neurodegeneration, sirtuin-1 (SIRT1), and mitochondrial dynamic protein expression in a mouse model of transient global cerebral ischemia and in vitro in primary mouse astrocyte cultures after simulated ischemia. Mice were subjected to 10 min bilateral common carotid artery occlusion plus hypotension with 5% isoflurane. After 2 h recovery mice were treated with intravenous injection of either anti-miR-200c or mismatch control. Memory function was assessed by Barnes maze at post-injury days 3 and 7. Mice were sacrificed at post-injury day 7 for assessment of brain cell-type specific expression of miR-200c, SIRT1, and the mitochondrial fusion proteins mitofusin-2 (MFN2) and OPA1 via complexed fluorescent in situ hybridization and fluorescent immunohistochemistry. Global cerebral ischemia induced significant loss of CA1 neurons, impaired memory performance and decreased expression of CA1 SIRT1, MFN2, and OPA1. Post-injury treatment with anti-miR-200c significantly improved survival, prevented CA1 neuronal loss, improved post-injury performance in Barnes maze, and was associated with increased post-injury expression of CA1 SIRT1 and MFN2 in astrocytes. In vitro, primary mouse astrocyte cultures pre-treated with miR-200c inhibitor prior to oxygen/glucose deprivation preserved expression of SIRT1 and MFN2, and decreased reactive oxygen species generation, whereas pre-treatment with miR-200c mimic had opposite effects that could be reversed by co-treatment with SIRT1 activator. These results suggest that miR-200c regulates astrocyte mitochondrial homeostasis via targeting SIRT1, and that CA1 astrocyte mitochondria and SIRT1 represent potential post-injury therapeutic targets to preserve cognitive function in survivors of global cerebral ischemia.

记忆障碍仍然是全球脑缺血幸存者的领先残疾，这是继发于海马氨基氨基-1（CA1）神经元神经退行性延迟的继发的。在缺血应力之后，诱导microRNA-200C（miR-200c），我们先前证明，抗MIR-200C的预处理可防止小鼠中的栓塞中风。在本研究中，我们评估了miR-200c在CA1神经变性，SIRTUIN-1（SIRT1）和线粒体动态蛋白表达中的作用在瞬时全球脑缺血的小鼠模型中和体外在原发性小鼠星形胶质细胞培养物中的作用。对小鼠进行10分钟双侧颈动脉闭塞和5％异氟烷的低血压。 2小时后，恢复后的小鼠用静脉注射抗MIR-200C或不匹配对照。在伤害后第3天和第7天通过巴恩斯迷宫评估记忆功能。在伤害后第7天处死小鼠，以评估miR-200C，SIRT1和线粒体融合蛋白miRofusin-2（MiR-200C，SIRT1）的特异性表达（ MFN2）和OPA1通过综合荧光原位杂交和荧光免疫组织化学。全球脑缺血引起了CA1神经元的显着丧失，记忆性能受损以及CA1 SIRT1，MFN2和OPA1的表达降低。抗MIR-200C的伤害后治疗可显着提高生存率，防止CA1神经元丧失，改善Barnes Maze的伤害后表现，并且与星形胶质细胞中CA1 SIRT1和MFN2的伤害后表达相关。在体外，原代小鼠星形胶质细胞培养物在氧/葡萄糖剥夺之前保留了SIRT1和MFN2的表达，并降低活性氧的产生，而用miR-200C进行预处理可能是相反的作用，可能是可能的相反作用通过与SIRT1激活剂共同治疗相反。这些结果表明，miR-200c通过靶向SIRT1调节星形胶质细胞线粒体稳态，并且CA1星形胶质细胞线粒体和SIRT1代表了潜在的伤害后治疗靶标，可以在全球脑缺血的幸存者中保留认知功能。