微小RNA-137调控帕金森致病基因参与癫痫发生的作用机制

Mechanism of miRNA-137 underlying epilepsy by regulating PD pathogenic gene

目的初步探究微小RNA-137(miR-137)调控帕金森致病基因(Parkin)发挥抗癫痫机制。方法选择SD健康清洁级雄性大鼠60只建立癫痫模型后随机分为模型组、低表达组、阴性组、3-MA组、联合组(n=12),分别给予生理盐水、miR-137低表达溶液、miR-137阴性对照溶液、自噬抑制剂3-MA溶液、miR-137拮抗剂联合3-MA溶液。另取12只大鼠为对照组,给予生理盐水。检测海马组织miR-137、微管相关蛋白轻链3-Ⅱ(LC3-Ⅱ)、线粒体自噬受体蛋白(NIX)、半胱氨酸天冬氨酸蛋白酶3(Capase-3)表达水平。结果与对照组比较,其他5组大鼠海马组织miR-137、Racine评分明显升高,模型组神经元凋亡率、阳性线粒体数目、LC3-Ⅱ、NIX、Capase-3表达明显升高(P<0.05)。与模型组比较,低表达组miR-137表达、Racine评分、神经元凋亡率、Capase-3降低,阳性线粒体数目、LC3-Ⅱ、NIX升高(P<0.05),3-MA组Racine评分、神经元凋亡率、Capase-3明显升高,阳性线粒体数目、LC3-Ⅱ、NIX表达明显降低(P<0.05)。联合组Racine评分、神经元凋亡率、Capase-3表达明显高于低表达组[(43.61±2.23)分vs(18.28±1.11)分,(25.68±1.29)%vs(13.34±1.24)%,1.96±0.19 vs 1.44±0.15,P<0.05];联合组阳性线粒体数目、LC3-Ⅱ、NIX明显低于低表达组(P<0.05)。结论下调miR-137可促进Parkin转位入线粒体,促进线粒体自噬,发挥神经元保护作用,并减轻癫痫症状。

Objective To study the mechanism of miR-137 underlying epilepsy by regulating Parkin gene.Methods Sixty healthy and clean male SD rats were randomly divided into model group,low expression group,negative group,3-MA group and combination group(n=12)after the epilepsy model of rats was established.The animals in different groups were given normal saline,low expression solution of miR-137,negative control solution of miR-137,3-MA solution,combined miR-137,LC3-Ⅱ,NIX,Capase-3 antagonists and 3-MA solution respectively.Another 12 rats served as a control group and were given normal saline.The expressions of miR-137,LC3-Ⅱ,NIX and Capase-3 in hippocampal tissues were detected by RT-qPCR and Western blot respectively.Results The Racine score,apoptosis rate of neurons,and expression levels of miR-137,LC3-Ⅱ,NIX,Capase-3 were significantly higher and the number of positive mitochondria was significantly greater in model group than in control group(P<0.05).The Racine score,apoptosis rate of neurons,expression levels of miR-137 and Capase-3 were significantly lower while the number of positive mitochondria was significantly greater and the expression levels of LC3-Ⅱand NIX were significantly higher in low expression group than in model group(P<0.05).The Racine score,apoptosis rate of neurons,expression level of Capase-3 were significantly higher while the number of positive mitochondria was significantly smaller and expression levels of LC3-Ⅱand NIX were significantly lower in 3-MA group than in model group(P<0.05).The Racine score,apoptosis rate of neurons,expression level of Capase-3 were significantly higher(43.61±2.23 vs 18.28±1.11,25.68%±1.29%vs 13.34%±1.24%,1.96±0.19 vs 1.44±0.15,P<0.05)while the number of positive mitochondria was significantly smaller and expression levels of LC3-Ⅱand NIX were significantly lower in combination group than in low expression group(P<0.05).Conclusion Down-regulated miR-137 can promote the translocation of Parkin into mitochondria and mitochondrial autophagy,protect neurons and improve symptoms of epilepsy.