Wang Dong, Li Zhenlu, Zhang Yukun, Wang Guangzhi, Wei Minghai, Hu Yan, Ma Shuo, Jiang Yue, Che Ningwei, Wang Xiaofeng, Yao Jihong, Yin Jian
Department of Neurosurgery, Second Hospital of Dalian Medical University, Dalian, Liaoning, China.
Department of Pharmacology, Dalian Medical University, Dalian, Liaoning, China.
Epilepsia. 2016 May;57(5):706-16. doi: 10.1111/epi.13348. Epub 2016 Mar 6.
MicroRNAs (miRNAs) are noncoding small RNAs that control gene expression at the posttranscriptional level. Some dysregulated miRNAs have been shown to play important roles in epileptogenesis. The aim of this study was to determine if miR-199a-5p regulates seizures and seizure damage by targeting the antiapoptotic protein silent information regulator 1 (SIRT1).
Hippocampal expression levels of miR-199a-5p, SIRT1, and acetylated p53 were quantified by quantitative real-time polymerase chain reaction (RT-PCR) and Western blotting in the acute, latent, and chronic stages of epilepsy in a rat lithium-pilocarpine epilepsy model. Silencing of miR-199a-5p expression in vivo was achieved by intracerebroventricular injection of antagomirs. The effects of targeting miR-199a-5p and SIRT1 protein on seizure and epileptic damage post-status epilepticus were assessed by electroencephalography (EEG) and immunohistochemistry, respectively.
miR-199a-5p expression was up-regulated, SIRT1 levels were decreased, and neuron loss and apoptosis were induced in epilepsy model rats compared with normal controls, as determined by up-regulation of acetylated p53 and cleaved caspase-3 expression. In vivo knockdown of miR-199a-5p by an antagomir alleviated the seizure-like EEG findings and protected against neuron damage, in accordance with up-regulation of SIRT1 and subsequent deacetylation of p53. Furthermore, the seizure-suppressing effect of the antagomir was partly SIRT1 dependent.
The results of this study suggest that silencing of miR-199a-5p exerts a seizure-suppressing effect in rats, and that SIRT1 is a direct target of miR-199a-5p in the hippocampus. The effect of miR-199a-5p on seizures and seizure damage is mediated via down-regulation of SIRT1. The miR-199a-5p/SIRT1 pathway may thus represent a potential target for the prevention and treatment of epilepsy and epileptic damage.
微小RNA（miRNA）是一类非编码小RNA，可在转录后水平调控基因表达。一些失调的miRNA已被证明在癫痫发生过程中起重要作用。本研究的目的是确定miR-199a-5p是否通过靶向抗凋亡蛋白沉默信息调节因子1（SIRT1）来调节癫痫发作和癫痫损伤。
在大鼠锂-匹罗卡品癫痫模型的癫痫急性、潜伏期和慢性期，通过定量实时聚合酶链反应（RT-PCR）和蛋白质印迹法对海马中miR-199a-5p、SIRT1和乙酰化p53的表达水平进行定量分析。通过脑室内注射抗miR来实现体内miR-199a-5p表达的沉默。分别通过脑电图（EEG）和免疫组织化学评估靶向miR-199a-5p和SIRT1蛋白对癫痫持续状态后癫痫发作和癫痫损伤的影响。
与正常对照组相比，癫痫模型大鼠中miR-199a-5p表达上调，SIRT1水平降低，并且通过乙酰化p53和裂解的半胱天冬酶-3表达的上调确定诱导了神经元丢失和凋亡。抗miR在体内敲低miR-199a-5p可减轻癫痫样EEG表现并防止神经元损伤，这与SIRT1的上调以及随后p53的去乙酰化一致。此外，抗miR的癫痫抑制作用部分依赖于SIRT1。
本研究结果表明，沉默miR-199a-5p在大鼠中具有癫痫抑制作用，并且SIRT1是海马中miR-199a-5p的直接靶点。miR-199a-5p对癫痫发作和癫痫损伤的影响是通过下调SIRT1介导的。因此，miR-199a-5p/SIRT1途径可能代表预防和治疗癫痫及癫痫损伤的潜在靶点。
