摘要
目的探讨缺氧缺血性脑损伤(HIBD)大鼠神经干细胞(NSCs)修复损伤的可能机制。方法传至2、3代的SD大鼠NSCs随机为空白对照组(未转染质粒者,CON),转染阴性对照质粒组(ncNSCs)和转染β-catenin siRNA真核表达质粒组(siNSCs),分别在不同脑组织匀浆上清液中培养,以模拟HIBD及正常脑内微环境。应用免疫荧光方法观察各组NSCs的分化情况;应用反转录PCR、Western blot法检测NSCs Ngn1,BMP4基因表达情况。结果与CON组比较,ncNSCs分化为神经元和少突胶质细胞的百分比明显增加(Pa<0.05),其中HIBD组增加较多;ncNSCs分化为星形胶质细胞的百分比显著减少(P<0.05)。与CON组相比,siN-SCs分化为神经元的百分比显著减少(P<0.01),分化为星形胶质细胞的百分比显著增加(P<0.01);少突胶质细胞的分化增加,但少于ncNSCs组(P<0.01),其中HIBD组分化为神经元和少突胶质细胞较多。与CON组比较,ncNSCs Ngn1 mRNA和Ngn1蛋白的表达显著增加(Pa<0.01),而BMP4 mRNA和BMP4蛋白的表达均显著减少(Pa<0.01);与CON组和ncNSCs组比较,siNSCs的Ngn1 mRNA和Ngn1蛋白表达显著减少(Pa<0.01),BMP4 mRNA和BMP4蛋白的表达显著增加(Pa<0.01),2个siNSCs组间Ngn1和BMP4的表达差异无统计学意义。结论 HIBD时受损的脑组织可进行自主修复,与β-catenin促进ncNSCs向神经元分化有关,BMP4和Ngn1在HIBD大鼠NSCs的增殖分化中起重要的协调作用。
Objective To investigate the roles of β - catenin in hypoxic - ischemic brain damage （HIBD） induced neurogenesis. Metho- ds Rats NSCs were divided into 5 groups randomized, then cultured in the supernatant of brain homogenate to simulate micro - environment of HIBD and normal brain. Immunocytochemical staining was performed simultaneously on the 5 groups of NSCs on precoated chamber slides to detect the differentiation of NSCs. Quantitative reverse transcription polymerase chain reaction was used to detect the relative contents of Ngnl mRNA and BMP4 mRNA in the 5 groups of NSCs. Western blot was used to detect the relative contents of Ngnl protein and BMP4 pro- tein in the 5 groups NSCs. Results HIBD or normal brain tissue extract cultures promoted ncNSCs differentiate into neurons and oligodendro- cytes but depressed astrogliosis, the effects of HIBD brain extract cultures was superior to the latter. β - catenin siRNA decreased the NSE - positive neurons and increased GFAP - positive astrocytes in the siNSCs in vitro. HIBD and normal brain tissue extract cultures increased the levels of Ngnl mRNA and decreased BMP4 mRNA of ncNSCs, the effect of HIBD brain extract cultures was superior to the latter; transfection of β - catenin siRNA could down - regulate the expression of Ngnl mRNA and up - regulate BMP4 mRNA of NSCs in vitro, respectively. HIBD and normal brain extract cuhures increased the levels of Ngnl protein and decreased BMP4 protein of ncNSCs, and the effects of HIBD brain extract cultures was superior to the latter;transfection of β - catenin siRNA could down - regulate the expression of Ngnl protein and up - regulate BMP4 protein of NSCs in vitro, respectively. Conclusions The signaling activity of β- catenin may control the production of newborn neurons and thus regulate the autonomous repair in the ischemic region after HIBD ;there is potential cooperative actions of BMP4 and Ngnl on differentiating rat NSCs in HIBD.
出处
《实用儿科临床杂志》
CAS
CSCD
北大核心
2012年第19期1511-1514,1517,共5页
Journal of Applied Clinical Pediatrics
基金
新疆维吾尔自治区高等学校科研计划项目(XJEDU2010S22)
