缝隙连接蛋白36在癫痫中的作用

发布时间：2018-05-17 13:32

本文选题：缝隙连接蛋白 + 癫痫　；参考：《吉林大学》2014年硕士论文

【摘要】：目的：癫痫是一种由多种病因引起的慢性脑部疾患，以神经元过度同步放电导致反复、发作性和短暂性的中枢神经系统功能失常为特征。缝隙连接在癫痫发病过程中发挥重要作用。本实验拟在建立PTZ诱导的癫痫大鼠模型的基础上，用缝隙连接通道阻断剂CBX进行干预处理，观察记录各组大鼠行为学、脑电图变化，并检测痫性发作后不同时间点Cx36及Caspase-3、Bcl-2、Bax的表达变化，进一步探讨缝隙连接蛋白在癫痫发作中的作用。方法：以大鼠腹腔内注射戊四氮制备急性癫痫模型为研究对象，将大鼠随机分为空白对照组、PTZ组、CBX组。采用HE染色、免疫组织化学染色、Western blot法等测定Cx36在痫性发作后不同时点(2h、4h、8h、12h、24h)的表达的变化，并用上述方法检测致痫24h后Caspase-3、Bcl-2、Bax在三组间表达的差异以及DAPI染色检测细胞凋亡。结果： 1、PTZ组和CBX组大鼠造模成功，脑电图均显示异常，出现棘波、多棘、多尖、尖-慢综合波等痫性电波。 2、免疫组织化学染色显示，大鼠皮质以及海马区均可以见到大量Cx36免疫阳性细胞，免疫反应产物主要分布于神经元的胞浆内。Cx36表达变化以海马CA1和CA3区为明显，Cx36蛋白表达在PTZ和CBX组均从致痫后2h开始增多，并持续至24h。PTZ组和CBX组分别与对照组同一时间点比较，海马CA1、CA3区Cx36表达增多具有统计学意义，CBX组与PTZ组各时间比较，无显著差异。 3、Western blot检测结果显示，Cx36蛋白表达在PTZ和CBX组均从致痫后2h开始增多，并持续至24h，与免疫组织化学染色结果一致。PTZ组和CBX组大鼠在致痫24h后，，Caspase-3，Bax蛋白表达增高，Bcl-2蛋白表达降低，CBX组与PTZ组比较，Caspase-3，Bax蛋白表达明显减少，Bcl-2蛋白表达较多。结论： 1． PTZ诱导的急性癫痫大鼠模型中，对照组模型组的Cx36在皮质以及海马区均大量表达。模型组的Cx36的表达强度明显高于对照组。PTZ组与CBX组间表达无差异。 2． Cx36表达及时程变化以海马CA1和CA3区为明显，在PTZ致痫后2h开始增多，并持续增多至24h。 3． PTZ诱导的急性癫痫大鼠模型出现神经元凋亡，CBX阻断缝隙连接通道的功能，减轻癫痫导致的神经元凋亡的出现。
[Abstract]:Objective: Epilepsy is a kind of chronic brain disease caused by many causes. It is characterized by recurrent, paroxysmal and transient central nervous system dysfunction caused by excessive synchronous firing of neurons. Gap junction plays an important role in the pathogenesis of epilepsy. On the basis of establishing epileptic rat model induced by PTZ, we used gap junction channel blocker CBX to intervene, observe and record the behavior and EEG changes of rats in each group. The expression of Cx36 and Caspase-3 / Bcl-2P Bax were detected at different time points after epileptic seizure to further explore the role of gap junction protein in epileptic seizures. Methods: The acute epilepsy model was established by intraperitoneal injection of pentylenetetrazol in rats. The rats were randomly divided into control group (PTZ) group and CBX group. The expression of Cx36 was detected by HE staining and immunohistochemical staining (Western blot). The expression of Caspase-3, Bcl-2Bcl-2Bax and apoptosis were detected by DAPI staining in 24 hours after epileptic seizure. Results: 1the rats in PTZ group and CBX group were successful in modeling, EEG showed abnormal, and epileptic waves such as spike wave, multi-spine wave, multi-tip wave and cusp-slow complex wave appeared. 2Immunohistochemical staining showed that a large number of Cx36 immunoreactive cells could be seen in the cortex and hippocampus of rats. Immunoreactive products mainly distributed in the cytoplasm of neurons. The expression of Cx36 protein in hippocampal CA1 and CA3 was significantly increased in both PTZ and CBX groups from 2 h after epilepsy, and continued to be compared between 24h.PTZ group and CBX group at the same time point as control group. There was no significant difference in the expression of Cx36 between the PTZ group and the hippocampal CA1 + CA3 group. 3Western blot analysis showed that the expression of Cx36 protein increased in both PTZ and CBX groups from 2 h after epilepsy. The results of immunohistochemical staining were consistent with the results of immunohistochemical staining for 24 h. The expression of Caspase-3 and Bax protein in CBX group was increased 24 hours after epilepsy. The expression of Bcl 2 protein in PTZ group was significantly lower than that in PTZ group, and the expression of Bcl 2 protein was significantly decreased in PTZ group. Conclusion: 1. In the PTZ induced acute epilepsy rat model, the Cx36 in the control group was highly expressed in the cortex and hippocampus. The expression of Cx36 in model group was significantly higher than that in control group and CBX group. 2. The changes of Cx36 expression were observed in the CA1 and CA3 regions of hippocampus, and increased at 2 h after PTZ induced epilepsy, and continued to increase to 24 h after PTZ induced epilepsy. 3. PTZ induced acute epileptic rat model showed the function of neuronal apoptosis and blocked gap junction channel, which alleviated the neuronal apoptosis induced by epilepsy.
【学位授予单位】：吉林大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R742.1

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