氟桂利嗪对苯妥英钠耐药癫小鼠的逆转作用

Reversion of resistance by flunarizine in phenytoin sodium-resistant epileptic mice

目的:研究氟桂利嗪对苯妥英钠耐药癫(drug resistant epilepsy)小鼠癫发作的逆转作用,初步探讨其作用机制。方法:采用戊四氮制作小鼠癫模型,灌胃给予苯妥英钠40mg/(kg·d),共14d,制作耐药癫小鼠模型。造模成功的耐药癫小鼠30只,按随机数字表法分为3组,分别为模型组、维拉帕米组和氟桂利嗪组。在给予苯妥英钠后30min,分别腹腔注射生理盐水、维拉帕米20mg/(kg·d)和氟桂利嗪8mg/(kg·d),连续给药4d,在第4天(d 4)观察给药后30min内小鼠癫发作的严重程度。然后处死小鼠,取脑。采用蛋白质印迹法(Western blot)检测P-糖蛋白(P-gp)表达情况;用免疫组化法测定半胱氨酸蛋白酶-3(caspase-3)的表达情况。结果:给药d 4,氟桂利嗪组小鼠癫发作严重程度较模型组显著减轻(P<0.01)。维拉帕米组和氟桂利嗪组小鼠脑内P-gp的表达量显著减少(P<0.01),且氟桂利嗪的效果优于维拉帕米(P<0.05);维拉帕米组和氟桂利嗪组小鼠皮层和海马半胱氨酸蛋白酶-3表达量也显著减少(P<0.01),氟桂利嗪的作用优于维拉帕米(P<0.05)。结论:氟桂利嗪能够有效降低苯妥英钠耐药小鼠的癫发作严重程度,其主要机制可能与降低小鼠脑组织P-gp的表达有关,同时氟桂利嗪也能降低耐药小鼠脑内半胱氨酸蛋白酶-3的表达,对脑组织可能有保护作用。

Objective：To investigate the reversion of resistance by flunarizine in phenytoin sodiurn resistant epileptic mice and explore its mechanism.Methods：A mouse model of phenytoin sodium-resistant epilepsy was established by gavage of phenytoin sodium 40 mg/（kg · d） for 14 days in pentylenetetrazole-kindled model of epilepsy in mice.By using random cluster method,30 mice with epilepsy resistant to phenytoin sodium were divided into 3 groups：the model group,the verapamil group and the flunarizine group.Thirty minutes after gavage of phenytoin sodium,normal saline,verapamil 20 mg/（kg · d） and flunarizine 8 mg/（kg · d） were administered ip to the mice,for a succession of 4 days.On d 4,the severity of epilepsy seizure was observed 30 minutes after medication.Then,the mice were sacrificed and brain samples were taken.Western blot was used to detect P-glycoprotein （P-gp） expression in the brain tissue,and immunohistochemistry was used to monitor the expression of caspase-3 in the brain tissue of mice.Results：After 4 consecutive days of medication,the severity of epilepsy seizure for the mice in the flunarizine group was significantly lighter than that of the model group （P＜0.01）.The expression of P-gp in the brain tissue for the mice in the flunarizine and verapamil groups decreased significantly （P＜0.01）,with the effect of flunarizine being superior than that of verapamil （P＜0.05）.The expressions of caspase-3 in cortex and hippocampus for the mice in the verapamil and flunarizine groups also decreased significantly （P＜0.01）,with the effect of flunarizine being superior than that of verapamil （P＜0.05）.Conclusion：Flunarizine could effectively alleviate the severity of epilepsy seizure in phenytoin sodium-resistant epileptic mice.The mechanism of which might be correlated with the reduction of P-gp expression in the brain tissue.Likewise,flunarizine could also suppress the expression of caspase-3 in the brain tissue of drug-resistant mice,which might be helpful to the protection of the brain tissue.