Lingo-1介导Rho-ROCK通路调控神经干细胞的分化  

Lingo-1 regulates neural stem cell differentiation via Rho-ROCK signaling pathway

作　　者：彭志明 赵晓阳[1] 朱凯 万勇[1] Peng Zhi-ming;Zhao Xiao-yang;Zhu Kai;Wan Yong(Department of Spine Surgery,the First Affiliated Hospital of Sun Yat-sen University,Guangzhou 510080,Guangdong Province,China)

机构地区：[1]中山大学附属第一医院脊柱外科,广东省广州市510080

出　　处：《中国组织工程研究》2018年第29期4669-4674,共6页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金(面上项目)(81472069;81171711)~~

摘　　要：背景:Lingo-1在神经元轴突再生、髓鞘形成等过程中具有重要作用,但其是否参与对神经干细胞分化的调控以及具体机制尚无定论。目的:观察干预神经干细胞Lingo-1表达水平对其分化方向的影响,并探索其可能机制。方法:从新生大鼠脑组织分离培养神经干细胞,经神经干细胞标志物检测鉴定后分别使用Lingo-1 shRNA慢病毒载体及阴性对照病毒转染神经干细胞,再用成神经元诱导分化培养基培养7 d,随后运用免疫荧光、免疫RhoA印迹技术检测神经元相关标志物表达,并检测、ROCK1蛋白表达水平。结果与结论:(1)从新生大鼠脑组织所分离培养的神经干细胞呈浮球状生长,高表达神经干细胞特异性标志物Nestin、Sox2与Pax6,符合神经干细胞特征;(2)慢病毒载体转染可在mRNA及蛋白质水平实现对神经干细胞Lingo-1的干扰,经诱导培养7 d后Lingo-1敲低的神经干细胞分化为神经元的比例显著增加,且伴随着RhoA、ROCK1蛋白表达水平的下降;(3)结果表明,Lingo-1参与对神经干细胞分化方向的调控,抑制Lingo-1可促进其向神经元方向分化,该过程可能与调控Rho-ROCK通路有关。BACKGROUND:It has been reported that Lingo-1 plays an important role in neuronal axon regeneration and myelination,but whether it participates in the regulation of neural stem cell differentiation remains unknown.OBJECTIVE:To examine the effect of Lingo-1 knockdown on the differentiation of neural stem cells and to explore its potential mechanism.METHODS:Neural stem cells were isolated from neonatal rat brain tissue and identified by determination of several neural stem cell specific markers.Then the cells were transfected with Lingo-1 shRNA lentivirus or negative control vectors,and were further cultured in induced differentiation medium for 7 days.Immunofluorescence and western blot techniques were thereafter used to detect the expression of neuron specific markers and to examine expression levels of RhoA and ROCK1.RESULTS AND CONCLUSION:The cells isolated from brain tissue of neonatal rats aggregated to form non-adherent spherical clusters with high expression of Nestin,Sox2,and Pax6,specific markers of neural stem cells,which exhibited the characteristics of neural stem cells.After transfected with lentivirus,expression of Lingo-1 was successfully knocked down at mRNA and protein levels.After 7 days of induction,expression of neuron specific markers was significantly increased in Lingo-1 knocked-down neural stem cells.Meanwhile,expressions of RhoA and ROCK1 were decreased.Thus,Lingo-1 can be involved in the differentiation of neural stem cells.Inhibition of Lingo-1 can induce neural stem cells to differentiate into neurons,which is possibly related to the regulation of Rho-ROCK signaling pathway.

关 键 词：神经干细胞 巢蛋白 β-微管蛋白Ⅲ 微管相关蛋白2 Rho/ROCK通路 Lingo-1蛋白 细胞分化 干细胞 国家自然科学基金 神经干细胞 细胞分化 神经元 组织工程 

分 类 号：R394.2[医药卫生—医学遗传学]