抑制miR-9通过上调FoxG1改善颞叶癫痫中Wnt/β-catenin通路异常导致的学习记忆障碍

Inhibition of miR-9 improving learning and memory dysfunction induced by Wnt/β-catenin pathway abnormal in temporal epilepsy by up-regulating FoxG1

目的探究Wnt/β-catenin通路在小鼠颞叶癫痫疾病中发生和发展的表达变化及抑制miR-9的表达通过上调FoxG1改善该通路异常导致的学习记忆障碍的分子机制。方法取健康C57BL/6小鼠随机分为对照组(n=3)和实验组(n=15),腹腔注射海人藻酸造癫痫模型,在癫痫持续状态发作(SE)1 h后终止,分别造模于1、7、14、30、60 d,观察癫痫反应并记录分级;实时荧光定量PCR检测海人藻酸诱导颞叶癫痫不同时间点小鼠海马组织中FoxG1、Wnt/β-catenin的表达水平;利用miRNA海绵技术将miR-9 sponge包装进腺相关病毒内(AAV-CamkⅡ-miR-9-sponge-EGFP)并注射到小鼠海马DG区,颞叶癫痫造模后记录分级并检测FoxG1、Wnt/β-catenin的表达情况;将AAV-CamkⅡ-miR-9-sponge-EGFP注射到小鼠海马DG区,颞叶癫痫造模后水迷宫和条件恐惧记忆检测小鼠的记忆情况。结果Wnt/β-catenin在不同时间点颞叶癫痫模型中较对照组表达下调,在14 d达到低峰值;FoxG1在不同时间点颞叶癫痫模型中表达情况与Wnt/β-catenin的表达呈正相关;抑制miR-9通过上调FoxG1改善了癫痫模型中Wnt/β-catenin通路异常从而改善了癫痫小鼠的学习记忆功能障碍。结论抑制miR-9的表达可通过上调FoxG1的表达,改善癫痫模型中Wnt/β-catenin通路异常,改善小鼠的学习记忆功能障碍,有希望成为颞叶癫痫治疗的潜在靶点。

Objective To investigate the expressions of Wnt/β-catenin in kainic acid-induced epilepsy in mice and the protective role of miR-9 in learning and memory dysfunction in kainic acid-induced epilepsy by regulating FoxG1.Methods C57/B6 mice were randomly divided into 5 groups(1,7,14,30,60 days).Temporal lobe epilepsy(TLE)model was induced by intraperitoneal injection of kainic acid.The model was terminated after 1 hour seizure of status epilepticus(SE),and the epilepsy response was observed and graded at 1,7,14,30 and 60 days respectively.Real-time fluorescence quantitative PCR was used to detect the expression levels of FoxG1 and Wnt/β-catenin in hippocampus of mice at different time points of TLE induced by kainic acid(1,7,14,30,60 days).The miR-9 sponge was packaged into adeno-associated virus and injected into DG area of hippocampus of mice.The expression of FoxG1 and Wnt/β-catenin was detected after TLE model was established.Learning and memory ability was detected by Morris Water Maze and Fear Conditioning Test.Results Wnt/β-catenin was down-regulated and FoxG1 was also down-regulated in epileptic models at different time points(P<0.01)and peaked at the 14th day.Inhibition expression of miR-9 increased the expression of FoxG1 and Wnt/β-catenin,alleviated epilepsy symptoms(P<0.01).Inhibition of miR-9 rescued the learning and memory dysfunction after TLE.Conclusions Inhibition of miR-9 could improve abnormal Wnt/β-catenin pathway induced learning and memory deficits in TLE by regulating FoxG1,which might become a potential target for TLE treatment.