摘要
目的观察大鼠脑缺血预处理后Notch信号通路对神经干细胞增殖分化的影响。方法将18只健康雄性SD大鼠随机分为假手术(Sham)组、缺血再灌注(MCAO)组、缺血预处理+缺血再灌注(CIP+MCAO)组。另将30只健康SD雄性大鼠随机分为假手术(Sham)组、缺血再灌注(MCAO)组、缺血预处理+缺血再灌注(CIP+MCAO)组、γ-分泌酶抑制剂+缺血再灌注(DAPT+MCAO)组、γ-分泌酶抑制剂+缺血预处理+缺血再灌注(DAPT+CIP+MCAO)组,采用线栓法制备MCAO模型,二次线栓法制备CIP+MCAO模型。通过Western blot检测缺血侧海马区Jagged1、Notch1、NICD、Hes1蛋白表达水平,采用Brdu/Nestin双免疫荧光标记法观察缺血侧海马区神经干细胞增殖分化的情况。结果缺血再灌注2 h Notch1的蛋白表达MCAO组明显高于Sham组(P <0. 05),CIP+MCAO组较MCAO组高(P <0. 05)。MCAO组在缺血再灌注24h Notch1、Jagged1、NICD、Hes1蛋白表达高于Sham组(P <0. 05),CIP+MCAO组较MCAO组低(P <0. 05)。加入DAPT后,Jagged1、Notch1、NICD、Hes1蛋白表达均降低(P <0. 05)。免疫荧光结果:MCAO组Brdu、Nestin荧光强度高于Sham组(P <0. 05)、CIP+MCAO组高于MCAO组(P <0. 05),在加入DAPT后,Brdu和Nestin值均降低(P <0. 05)。结论脑缺血预处理后使Notch1的表达高峰前移;脑缺血预处理能激活并上调Notch信号通路的表达、促进神经干细胞增殖分化而产生脑保护作用。
Objective To observe the effects of Notch signaling pathway on proliferation and differentiation of neural stem cells after cerebral ischemic preconditioning in rats. Methods 18 healthy male adult SD rats were randomly divided into Sham operation group（ Sham）,ischemia reperfusion group（ MCAO）,ischemic preconditioning ＋ ischemia reperfusion group（ CIP ＋ MCAO）. 30 healthy male adult SD rats were selected for experiment. The middle cerebral artery occlusion（ MCAO） model was prepared by suture-occluded method,and CIP plus MCAO model was prepared by twice occlusion with thread. The rats were randomly divided into sham operation group（ Sham）,ischemia reperfusion group（ MCAO）,cerebral ischemic preconditioning plus ischemia reperfusion group（ CIP ＋ MCAO）,Gamma secretase inhibitors plus ischemia reperfusion group（ DAPT ＋ MCAO）,and Gamma secretase inhibitors,cerebral ischemic preconditioning plus ischemia reperfusion group（ DAPT ＋ CIP ＋ MCAO）. The expression levels of Notch1,Jagged1,NICD and Hes1 in ischemic hippocampus were assessed through Western blot. The proliferation and differentiation of neural stem cells in ischemic hippocampus were observed by Brdu and Nestin double immunofluorescence labeling. Results Notch1 protein expression in Group CIP ＋ MCAO was higher than that in Group MCAO 2 hours after ischemia-reperfusion. The expression of Notch1,Jagged1,NICD,and Hes1 of MCAO group was significantly increased 24 hours after ischemia-reperfusion compared to the sham group（ P〈 0. 05）; it was significantly lower in Group CIP ＋ MCAO than that in Group MCAO（ P〈 0. 05）. Under DAPT conditions,the expression of Notch1,Jagged1,NICD and Hes1 in Group DAPT ＋ MCAO were significantly lower than those in Group MCAO; and they were significantly lower in Group DAPT ＋ CIP ＋ MCAO than those in Group CIP ＋ MCAO（ P〈 0. 05）. The fluorescence intensities of Brdu and Nestin in MCAO group were significantly increased 24 hours after ischemia-reperfusion compared to the sham group（ P〈 0. 05）,and they were significantly higher in Group CIP ＋ MCAO than those in Group MCAO（ P〈 0. 05）. Under DAPT conditions,the fluorescence intensities of Brd U and Nestin in Group DAPT ＋ MCAO were significantly lower than those in Group MCAO; they were significantly lower in Group DAPT ＋ CIP ＋ MCAO than those in Group CIP ＋ MCAO（ P〈 0. 05）. Conclusions Cerebral ischemic preconditioning advances the peak of Notch1 expression. Cerebral ischemic preconditioning activates and upregulates the expression of Notch signaling pathway,promotes the proliferation and differentiation of neural stem cells,and protects brain.
作者
蒋琼
陈丽
黄宽
王容
吴正华
JIANG Qiong, CHEN Li, HUANG Kuan , WANG Rong , WU Zheng - hua(Graduate School of Gannan Medical University, Ganzhou 541000, Jiangxi, Chin)
出处
《广东医学》
CAS
2018年第18期2718-2722,共5页
Guangdong Medical Journal
基金
国家自然科学基金资助项目(编号:81260202)
关键词
脑缺血预处理
NOTCH信号
神经干细胞
增殖分化
cerebral ischemic preconditioning
Notch signaling pathway
NSCs
proliferation
differentiation
