海马微量注射酪氨酸激酶受体结合蛋白B3激动剂对癫痫模型大鼠自发性痫性发作的影响及其机制  

Effect of microinjection of tyrosine kinase receptor binding protein B3 agonist into hippocampus on spontaneous seizures in epileptic model rats and its mechanisms

作　　者：刘田田 肖波[2] 张敏[3] 王欣格 刘恒方[3] 滕军放[1] Liu Tiantian;Xiao Bo;Zhang Min;Wang Xinge;Liu Hengfang;Teng Junfang(Department of Neurology,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China;Department of Neurology,Xiangya Hospital,Central South University,Changsha 410008,China;Department of Neurology,the Fifth Affiliated Hospital of Zhengzhou University,Zhengzhou 450054,China)

机构地区：[1]郑州大学第一附属医院神经内科,450052 [2]中南大学湘雅医院神经科,长沙410008 [3]郑州大学第五附属医院神经科,450054

出　　处：《中华行为医学与脑科学杂志》2021年第11期961-968,共8页Chinese Journal of Behavioral Medicine and Brain Science

基　　金：国家自然科学基金项目(81901326);河南省高等学校重点科研项目计划(19A320038)。

摘　　要：目的探讨海马内注射酪氨酸激酶受体结合蛋白B3(Ephrin-B3)激动剂对癫痫模型大鼠自发性痫性发作及海马组织分泌型糖蛋白Reelin及磷酸化接头蛋白(p-Dab1)表达的影响。方法将78只大鼠按照体质量匹配原则随机分成对照组和模型组,每组39只。对照组大鼠正常饲养,模型组大鼠采用氯化锂-匹罗卡品注射法建立大鼠癫痫模型,在癫痫持续状态诱导成功后的急性期(7 d),静止期(14 d)和慢性期(60 d)取海马组织,采用免疫组化及蛋白质免疫印迹法(Western blot)检测癫痫模型大鼠和正常大鼠海马组织Reelin蛋白及p-Dab1蛋白表达变化,每个时间点每组随机选取13只大鼠。另取48只大鼠,按照随机数字表法分为正常对照Fc-control组、正常对照EphB3-Fc组、癫痫Fc-control组和癫痫EphB3-Fc组,每组12只,前两组正常饲养,后两组进行癫痫造模,造模后7 d,所有大鼠按照分组分别进行双侧海马内注射Ephrin-B3的激动剂(EphB3-Fc)和Ephrin-B3激动剂的对照剂Fc-control,1次/d,连续7 d。给药结束后,观察两周内大鼠行为与脑电图的变化,采用免疫组化及Western blot检测Reelin蛋白及p-Dab1蛋白表达变化。采用SPSS 21.0进行统计分析,多组间比较采用单因素方差分析,两两比较采用Tukey's检验。结果免疫组化和Western blot结果均显示,模型组大鼠海马组织中Reelin和p-Dab1蛋白在致痫后7 d、14 d、60 d表达均低于对照组(均P<0.01)。免疫组化结果显示,与癫痫后Fc-control组相比,癫痫后EphB3-Fc组大鼠海马组织中p-Dab1蛋白的平均光密度值更高[(0.41±0.04),(0.58±0.06),P<0.05]。Western blot结果显示,癫痫后EphB3-Fc组大鼠海马组织中p-Dab1蛋白表达显著高于癫痫后Fc-control组[(1.34±0.04),(2.26±0.10),P<0.01]。动物行为学监测显示,与癫痫后Fc-control组相比,癫痫后EphB3-Fc组大鼠自发性痫性发作持续时间更短[(39.00±1.79)s,(26.50±1.87)s;t=23.21,P<0.01],频率更低[(2.00±0.89)次,(0.50±0.55)�Objective To investigate the effects of hippocampal injection of tyrosine kinase receptor binding protein B3(Ephrin-B3)agonist on spontaneous seizures and the expression of hippocampal secretory glycoprotein(Reelin)and phosphorylated adaptor protein(p-Dab1)in epileptic model rats.Methods Seventy-eight rats were randomly divided into control group and model group according to body mass matching with 39 rats in each group.The rats in control group were fed normaly,and the rats in model group were established epilepsy model by intraperitoneal injection of lithium chloride pilocarpine.The hippocampal tissues were taken in the acute phase(7 days),quiescent phase(14 days)and chronic phase(60 days)after the successful induction of status epilepticus.The levels of Reelin protein and p-Dab1 protein in the hippocampal tissues of epileptic model rats and normal rats were detected by immunohistochemistry and Western blot.And thirteen rats were randomly selected at each time point.Another 48 rats were randomly divided into normal Fc-control group,normal EphB3-Fc group,epilepsy Fc-control group and epilepsy EphB3-Fc group,with 12 rats in each group.Rats in the first two groups were fed normally,and those in the latter two groups were established epileptic model.Seven days after modeling,all rats were injected into bilateral hippocampus with EphB3-Fc(Ephrin-B3 agonist)and FC control(control agent of Ephrin-B3 agonist)according to the grouping,once a day for 7 days.After administration,the changes of behavior and EEG were observed within two weeks.At the same time,the expression of Reelin protein and p-Dab1 protein were detected by immunohistochemistry and Western blot.SPSS 21.0 was used for statistical analysis,One-way ANOVA was used for multi group comparison,and Tukey's test was used for pairwise comparison.Results The results of immunohistochemistry and Western blot showed that compared with the control group,the levels of Reelin and p-Dab1 protein in hippocampus of model group decreased significantly at 7,14 and 60 days aft

关 键 词：癫痫 海马齿状回 酪氨酸激酶受体结合蛋白B3 分泌型糖蛋白 磷酸化接头蛋白 自发性痫性发作 

分 类 号：R742.1[医药卫生—神经病学与精神病学]