MicroRNA-134/CREB/pCREB通路在癫痫中的表达及意义

目的探讨miR-134、CREB、pCREB在癫痫大鼠海马及难治性癫痫患者颞叶脑组织中的表达及意义。方法难治性癫痫患者及非癫痫对照组颞叶组织、氯化锂-匹罗卡品癫痫大鼠及空白对照组海马组织中,应用实时荧光定量PCR技术检测microRNA-134（miR-134）的表达,用Western blot方法检测CREB及p CREB的表达,用免疫组织化学方法检测人脑颞叶皮质及大鼠海马区CREB、p CREB的表达。结果与对照组相比miR-134表达在难治性癫痫患者中明显降低（P〈0.05）,在癫痫模型组中点燃后3、7、14、60 d明显降低（P〈0.05）,1 d与30 d表达降低较对照组差异无显著性（P〉0.05）;癫痫模型组CREB在3、7、14、60 d时间点明显升高（P〈0.05）、pCREB各时间点表达均高于空白对照组（P〈0.05）。结论难治性癫痫患者颞叶皮质及癫痫动物海马中miR-134表达下降,CREB、pCREB表达升高,提示其可能在癫痫发生发展机制中起重要作用。

Expression and significance of signaling pathway of miR-134/CREB/pCREB in patients with epilepsy and in epileptic rats

Objective To investigate the expression of microRNA-134 （ miR-134 ） , CREB and pCREB in the temporal lobe tissue of patients and epileptic rats and to explore their roles in pathogenesis of epilepsy.Methods Temporal lobe tissue samples of 14 patients with refractory epilepsy and 10 non-epileptic patients, and hippocampus and brain tissue samples of 42 rats were used in this study.Forty-two healthy adult male Sprague-Dawley rats were randomly divided into 6 epilepsy groups （24 h, 72 h, 7 d, 14 d, 30 d, and 60 d after kindling epilepsy） and a normal control group （n=6 for all groups） .The rat model of epilepsy was generated by intraperitoneal injection of 127 mg/kg lithium chloride and 16-20 h later, 35 mg/kg pilocarpine.In the temporal lobe tissue of patients and hippocampal tissue of rats, the expression level of miR-134 was detected by real-time polymerase chain reaction.The expression levels of CREB and pCREB were determined by Western blot, and CREB and pCREB localization was assessed by immunohistochemistry.Results Compared with the control rats, the expression of miR-134 was significantly decreased in the temporal lobe tissue of experimental rats at 72 h,7 d,14 d, 60 d after kindling （P〈0.05）,and no significant change at 24 h and 30 d after kindling （P〉0.05）. Expression of miR-134 in patients with refractory epilepsy was significantly lower than that of the controls （ P〈0.05 ） , while up-regulation of CREB expression was at the same time points （P〈0.05）.Up-regulation of pCREB expression was at all the time points after kindling （P〈0.05）.CREB and p-CREB expressions were seen in the nuclei of neurons, and significantly higher in patients with refractory epilepsy and epileptic rats.Conclusions The expression of miR-134 is significantly decreased and that of CREB and pCREB was significantly increased in the temporal lobe tissue of patients with refractory epilepsy and the hippocampal tissue of epileptic rats.These findings indicate that the signaling pathway of miR-134/C