戊四氮致痫大鼠脑N-cadherins和NT-4的变化及灵芝孢子粉和百里香精油的干预

【作者】 王芳芳；

【导师】 王淑秋； 马小茹； 刘君星；

【作者基本信息】 佳木斯大学 ， 病理学与病理生理学， 2010， 硕士

【摘要】 目的:观察和探讨戊四氮致痫大鼠脑N-cadherins和NT-4的变化及灵芝孢子粉和百里香精油的干预治疗,进而研究癫痫的发病机理及灵芝孢子粉和百里香精油的作用机制。方法:健康雄性Wistar实验大鼠50只,随机分为正常对照组、癫痫模型组、灵芝孢子粉治疗组、百里香精油治疗Ⅰ组、百里香精油治疗Ⅱ组,每组10只。模型组和治疗组均用致痫亚惊厥剂量的PTZ(35mg/kg)腹腔注射,每24小时1次,持续给药28天;正常对照组进行腹腔注射等容生理盐水。灵芝孢子粉组经灌胃方式给予灵芝孢子粉(30g/L),灌胃体积10.0ml/kg;百里香治疗组按不同浓度分为Ⅰ、Ⅱ两组,给予分别雾化吸入1﹪和2﹪百里香精油30分钟;模型组和正常组以等容生理盐水灌胃。每日监测并记录癫痫发作的级别、潜伏期及持续时间,给致痫药28天,停止用药7天,再一次应用相等剂量的PTZ测试,凡出现连续5次Ⅱ级以上痫性发作为达到点燃标准。在低温条件下迅速取脑,通过免疫组化和Western-Blotting检测皮质和海马区N-cadherins和NT-4的变化;采用电镜法检测海马神经元细胞的变化。结果:实验性癫痫大鼠模型制作成功,灵芝孢子粉组、百里香精油治疗Ⅰ组、百里香精油治疗Ⅱ组和癫痫模型组实验动物全部达到点燃标准,与癫痫模型组动物相比,灵芝孢子粉组和百里香精油治疗Ⅰ组、百里香精油治疗Ⅱ组大鼠潜伏期明显延长,但持续时间之间的差别没有显著性。免疫组化和Western-Blotting检测实验结果表明:癫痫模型组实验大鼠脑海马和皮质N-cadherins含量比正常对照组增高,差异具有显著性(P<0.01);灵芝孢子粉组N-cadherins含量比癫痫模型组下降,差异具有显著性(P<0.01)。百里香精油治疗Ⅰ、Ⅱ组N-cadherins含量均比癫痫模型组下降,差异具有显著性(P<0.01)。同时癫痫模型组大脑海马和皮质NT-4含量比正常对照组增加,差异具有显著性(P<0.05);灵芝孢子粉组NT-4含量比癫痫模型组增加,差异有显著性(P<0.01)。百里香精油治疗Ⅰ、Ⅱ组的NT-4含量比癫痫模型组增加,差异有显著性(P<0.05)。电镜检测癫痫模型组海马神经元改变,细胞线粒体增多,细胞核皱缩。在用灵芝孢子粉干预后,海马神经元无明显改变,百里香精油治疗Ⅰ、Ⅱ组干预后,海马神经元无明显改变。结论:灵芝孢子粉和百里香精油能够降低N-cadherins表达对癫痫大鼠中枢神经系统的毒性损害,调整病变神经元兴奋性,起到抗癫痫作用,还能增强NT-4的表达从而减轻癫痫发作给神经系统的损伤,对神经细胞起保护作用。更多还原

【Abstract】 Objective:To observe and discuss the influence of Ganoderma lucidum spores and Thymus mongolicus Ronn on the N-cadherins, NT-4 content in the brain of rat with epilepsy caused by Pentetrazole (PTZ) .And further study the epilepsy mechanism and anti-epilepsy mechanism of ganoderma lucidum spores and Thymus mongolicus Ronn. Methods:50 male wister rats (weighing 180-220g ,bought from Jiamusi university) were stochastic divided into five groups: (1) the control group(n = 10), in which rats received oral administration of the normal saline and intraperitoneal injections,(2)PTZ group (n = 10), in which rats lead to epilepsy were injected with a subconvulsant dose PTZ (35 mg/kg in 0.9% saline) intraperitoneally and received saline orally; (3) Ganoderma Lucidum spores group (n = 10), in which rats were injected intraperitoneally with PTZ (35 mg/kg in 0.9% saline) and received oral administration of Ganoderma Lucidum spores ( 300 mg/kg in normal saline ). (4) Thymus mongolicus RonnⅠgroup (n = 10), in which rats were injected intraperitoneally with PTZ (35 mg/kg in 0.9% saline) and received aerosol inhalation for 30 minutes. (5) Thymus mongolicus RonnⅡgroup (n = 10), in which rats were injected intraperitoneally with PTZ (35 mg/kg in 0.9% saline) and received aerosol inhalation for 30 minutes. 28 days later , withdraw drugs continuously for one week, then these rats received a final challenge dose of PTZ (35 mg/kg) to check the persistence of enhanced susceptibility to the convulsant. Only those PTZ-treated rats shows stageⅡor more than stageⅡconvulsion for at least five times continuously were used in the study. Control rats received injections of saline instead of the PTZ challenge dose. Put to death rats and get brain quickly at lower temperature, measure the content change of N-cadherins and NT-4 in cerebral cortex、hippocampus with immunohistochemical staining and western-blotting. observe influence on brain of epileptic rats in electron microscopy. Results:Rat model of experimental epilepsy were produced successfully .The animal of Garoderma lucidum spores groups, Thymus mongolicus RonnⅠ,Ⅱgroup and epileptic group all reach the "igniting" standard. Compared with epilepsy model group, the latent period of ganoderma lucidum spores group , Thymus mongolicus RonnⅠ,Ⅱgroup extended obviously, but duration had no obvious difference. Immunohistochemical staining and Western-Blotting revealed that the content of N-cadherins in cortex and hippocampus increased obviously in the epileptic model group as compared with the control group (P <0.01); the content of N-cadherins in the ganoderma lucidum spores group is lower than that in the epileptic model group (P<0.01); the content of N-cadherins in the Thymus mongolicus RonnⅠ,Ⅱgroup are lower than that in the epileptic model group (P<0.01). The content of NT-4 in cortex and hippocampus increased in the epileptic model group as compared with the control group (P<0.05); the content of NT-4 in the ganoderma lucidum spores group is higher than that in the epileptic model group (P<0.01). the content of NT-4 in the Thymus mongolicus RonnⅠ,Ⅱgroup are higher than that in the epileptic model group (P<0.05). Detected the influence on hippocampal neurons of epileptic model by transmission electron microscopy , what we found is that hippocampal neurons mitochondria increased, nucleus shrank. After the intervention of Ganoderma lucidum spore powder ,Thymus mongolicus RonnⅠ,Ⅱgroup, no significant changes happened in hippocampal neurons. Conclusions:Ganoderma lucidum spores powder and Thymus mongolicus Ronn can weaken the noxious injure for over expression of N-cadherins on centre nerve system of rat, regulation of neuronal excitability, prevent the damages of epilepsy to nerve cells; what is more, ganoderma lucidum spores powder and Thymus mongolicus Ronn can increase expression NT-4, so it seems to play a role in relieving seizure, protecting neurons.更多还原