细胞外信号调节激酶对细胞周期调控的影响  

Effect of Extracellular Signal-Regulated Kinases on the Cell Cycle Regulation After Focal Cerebral Ischemia

作　　者：张贵斌[1] 王伟[2] 徐运兰[2] 王萍[2] 田代实[2] 

机构地区：[1]湖北医药学院附属襄阳医院神经内科,湖北襄阳441000 [2]华中科技大学同济医学院附属同济医院神经内科,湖北武汉430030

出　　处：《武汉大学学报（医学版）》2011年第6期717-722,F0002,共7页Medical Journal of Wuhan University

基　　金：国家自然科学基金重点资助项目(编号:30230140)

摘　　要：目的:研究脑缺血后细胞外信号调节激酶(ERK)对细胞周期调控的影响。方法:建立光化学法诱导大鼠局灶性脑缺血模型,随机分为脑缺血组(对照组及干预组)和假手术组。干预组于缺血前30min尾静脉注入ERK阻滞剂U0126,对照组尾静脉注入相同体积不含U0126的DMSO稀释溶液。应用免疫组织荧光化学法观察细胞周期蛋白D1(Cyclin D1)和细胞周期蛋白E(Cyclin E)阳性细胞表达;免疫印迹(Western blot)检测磷酸化ERK1/2(pERK1/2)、Cyclin D1和Cyclin E的蛋白表达;半定量逆转录-聚合酶链反应(RT-PCR)观察转录因子E2FmRNA的表达。结果:干预组Cyclin D1和Cyclin E阳性表达的细胞数较对照组显著减少(P<0.05);缺血对照组pERK1/2蛋白表达明显强于干预组(P<0.05),4h时间点表达最为明显,12h时间点恢复到基线水平,Cyc-lin D1和Cyclin E表达6h开始升高,12h表达最为明显,干预组较对照组显著减弱(P<0.05);缺血对照组E2FmRNA的表达显著强于干预组和假手术组(P<0.05),以7d表达最为明显。结论:ERK在大鼠脑缺血中发挥重要作用,抑制脑缺血引起的pERK1/2磷酸化可降低细胞周期蛋白Cyclin D1、Cyclin E和E2F的表达,即细胞外信号调节激酶可影响细胞周期的调控。Objective： To investigate the effect of extracellular signal-regulated kinases（ERKs） on cell cycle regulation after ischemia.Methods： Cerebral ischemic model was induced by photochemistry method.Animals were divided randomly into cerebral ischemia groups（control and U0126 treated groups） and sham group.Rats in U0126 treated group were subjected to ERK inhibitor U0126 injection at 30 min pre-ischemia through caudal veins,while rats in control group were subjected to identical volume of DMSO solution without U0126.Positive immunostaining for Cyclin D1 and Cyclin E were detected by immunohistofluorescence method.Immunoblot showed protein expression of ERK1/2-phosphorylating,Cyclin D1,and Cyclin E in ischemic brain cortex.Semi-quantitation reverse transcription-polymerase chain reaction（RT-PCR） showed mRNA expression of transcription factor E2F in ischemic brain cortex.Results： Strong reduction of Cyclin D1-positive and Cyclin E-positive cells was presented after U0126 treatment as compared with those in the control.Western blot results showed pERK1/2-positive,Cyclin D1-positive,and Cyclin E-positive expression in control group was significantly increased after U0126 tveatment as compared with those in U0126 treated group,the strongest pERK1/2-positive expression was presented at 4 h post-ischemia and returned to baseline level at 12 h,and the strongest Cyclin D1-positive and Cyclin E-positive expression was presented at 12 h.RT-PCR showed strong increase of E2F mRNA in control group as compared with the treated and sham groups,and the strongest expression was presented at 7 d.Conclusion： ERK pathway plays a very important role in cerebral ischemia.Inhibiting ERK1/2 phosphorylation post-ischemia reduces the expression of Cyclin D1,Cyclin E and E2F,which indicates that ERK can affect cell cycle regulation.

关 键 词：ERKS U0126 CYCLIN D1 CYCLIN E E2F 细胞周期 脑缺血 

分 类 号：Q253[生物学—细胞生物学]