惊厥持续状态幼年大鼠海马中葡萄糖调节蛋白78与半胱氨酸门冬氨酸特异性蛋白酶12的表达及依达拉奉对其影响  

Effects of edravone on the expression of GRP78, Caspase-12, and neuron apoptosis in juvenile rat hippocampus after status convulsivus

作　　者：李光乾[1] 王海萍[1] 蒋春明[2] 

机构地区：[1]浙江中医药大学附属杭州第六医院杭州市儿童医院神经科,杭州310014 [2]杭州市第一人民医院儿科

出　　处：《中华儿科杂志》2011年第1期53-59,共7页Chinese Journal of Pediatrics

摘　　要：目的 观察幼年大鼠惊厥持续状态（status convulsion,SC）后脑组织海马中内质网分子伴侣葡萄糖调节蛋白78（glucose regulated protein,GRP78）、半胱氨酸门冬氨酸特异性蛋白酶12（Caspase-12）的表达及神经细胞凋亡的变化,并探讨依达拉奉（edaravone,ED）对三者的影响.方法 将195只SD幼年雄性大鼠随机分为生理盐水对照组（NS组）、惊厥持续状态组（SC组）和依达拉奉预处理组（ED组）,每组65只,各组均按SC后处死时间点分为4、12、24、48、72 h五个亚组,每组13只.采用氯化锂-匹鲁卡品化学点燃法制备幼年大鼠SC模型.应用逆转录多聚酶链反应（RT-PCR）法检测GRP78 mRNA和Caspase-12 mRNA的表达,免疫组化法检测大鼠海马中GRP78和Caspase-12 蛋白表达情况,TUNEL法检测神经细胞凋亡数的变化.结果 （1）免疫组化结果显示SC组幼年大鼠海马中GRP78（12 h组0.1480±0.0164,24 h组0.1682±0.0114,48 h组0.1540±0.0102）和Caspase-12（12 h组0.1325±0.0165,24 h组0.1794±0.0213,48 h组0.1525±0.0423,72 h组0.1309±0.0199）表达增强,与NS组（GRP78：12 h组0.1214±0.0147,24 h组0.1272±0.0177,48 h组0.1260±0.0157;Caspase-12：12 h组0.1050±0.0121,24 h组0.1041±0.0151,48 h组0.1058±0.0222,72 h组0.1036±0.0186）比较差异有统计学意义（P＜0.01或＜0.05）;与SC组比较,ED组GRP78（12 h组0.1550±0.0131,24 h组0.1886±0.0154,48 h组0.1721±0.0151）表达明显增强,而Caspase-12（12 h组0.1211±0.0184,24 h组0.1545±0.0205,72 h组0.1085±0.0219）表达明显降低,差异均有统计学意义（P＜0.01或P＜0.05）;（2）RT-PCR法检测结果显示GRP78 mRNA和Caspase-12 mRNA表达趋势与蛋白基本相似;（3）SC组在惊厥12 h（11.41±2.37）时间点海马CA1区TUNEL阳性细胞数已显著高于NS组（P＜0.01）,48 h（28.78±5.11）达峰,而ED组TUNEL阳性细胞数在12～72 h时间点（12 h组8.98±2.22,24 h组13.09±2.54,48 h组20.57±4.89）均较SC组显著下降（P＜0.01或P＜0.05）,但仍高Objective To observe the expression of GRP78 （glucose regulated protein, GRP78）,Caspase-12 and the change of neuron apoptosis in the juvenile rat hippocampus after status convulsivus （ SC）, and to explore the effect of edaravone on them. Methods One hundred and ninety-five juvenile male Sprague-Dawley（SD） rats were randomly divided into normal saline control group （NS group）, status convulsivus group（SC group）and edaravone treatment group（ ED group）. Each group was further divided into five subgroups in different executed time points after SC. The rats in status convulsivus group were kindled into epilepsy by lithium-pilocarpine method. Expression of GRP78 mRNA and caspase-12 mRNA was detected with reverse transcription-polymerase chain reaction （RT-PCR） method. Expressions of GRP78and caspase-12 protein were detected with immunohistochemical methods. The neuron gpoptosis was observed by TdT-mediated dUTP nick end labeling （TUNEL）. Results （1） Measured by immunohistochemistry the value of OD of GRP78 （ 0. 1480 ± 0. 0164,0. 1682 ± 0. 0114, and 0. 1540 ±0. 0102,respectively, 12 h-48 h points ） and caspase-12 （ 0. 1325 ± 0. 0165,0. 1794 ± 0. 0213,0. 1525 ±0. 0423, and 0. 1309 ± 0. 0199, respectively, 12 h-72 h points）positive cells in the SC group increased, there was a significant difference compared with NS group （GRP78： 0. 1214 ± 0. 0147,0. 1272 ± 0. 0177, and 0. 1260 ±0. 0157, respectively, 12 h-72 h points. Caspase-12：0. 1050 ±0. 0121,0. 1041 ± 0. 0151,0. 1058±0. 0222, and 0. 1036 ± 0. 0186, respectively, 12 h-72 h points ） （ P 〈 0. 01, or P 〈 0. 05 ）. By ED intervention GRP78 （ 0. 1550 ± 0. 0131, 0. 1886 ± 0. 0154, and 0. 1721 ± 0. 0151, respectively, 12 h-48 h points） positive cells value of the OD increased as compared with SC group （P 〈 0. 01, or P 〈 0. 05 ）. and caspase-12（0. 1211 ±0.0184,0. 1545 ±0.0205,and 0. 1085 ± 0.0219,respectively, 12 h,24 h and 72 h points） positive cells value of the A decreased as comp

关 键 词：发作 热休克蛋白质类 半胱氨酸天冬氨酸蛋白酶12 细胞凋亡 依达拉奉 

分 类 号：R720.597[医药卫生—急诊医学]