勿动蛋白A胞外肽残基1-40修饰神经干细胞促进轴突再生  

NogoA extracellular peptide residues 1-40 modified neural stem cells to promote axonal regeneration

作　　者：张弼 胡利红[2] 李旭生[3] 张宇飞 马俊驰 袁海峰[3] Zhang Bi;Hu Lihong;Li Xusheng;Zhang Yufei;Ma Junchi;Yuan Haifeng(Department of Anesthesiology,Ningbo Medical Center Li Huili Hospital,Ningbo 315048,China;Department of Geriatrics and Special Needs Medicine,General Hospital of Ningxia Medical University,Yinchuan 750003,China;Department of Spinal Department,General Hospital of Ningxia Medical University,Yinchuan 750003,China;The Second Department of Spine,Baoji Hospital of Traditional Chinese Medicine,Baoji 721001,China;Lanzhou Hospital of Traditional Chinese Medicine,Lanzhou 730050,China)

机构地区：[1]宁波市医疗中心李惠利医院麻醉科,宁波315048 [2]宁夏医科大学总医院老年与特需医学科,银川750003 [3]宁夏医科大学总医院脊柱骨科,银川750003 [4]宝鸡市中医医院脊柱二科,宝鸡721001 [5]甘肃省兰州市中医院创伤骨三科,兰州730050

出　　处：《中华实验外科杂志》2023年第9期1799-1803,共5页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金(81160220);宁夏回族自治区中央引导地方科技发展专项项目(2022FRD05038);宁夏医科大学暨临床医学院一级学科建设立项项目(NXYLXK2017A05)。

摘　　要：目的构建勿动蛋白A胞外肽残基1-40(NEP1-40)基因工程神经干细胞(NEP1-40-NSCs)并探究其对神经元轴突再生的影响。方法制备NEP1-40-NSCs,检测目的基因的表达量,同时鉴定其分化潜能;NEP1-40-NSCs组与神经干细胞(NSCs)组分别与大鼠海马组织神经元共培养后测量各组神经元轴突新增面积,同时鬼笔环肽染色观察生长锥及丝状伪足结构。多组间比较用单因素方差分析,两组间比较用t检验。结果NEP1-40-NSCs组β3-微管蛋白(Tuj-1)、少突胶质细胞转录因子2(Oligo2)及髓鞘碱性蛋白(MBP)所标记的阳性细胞数高于NSCs组(47.001±16.670比30.333±3.480、38.001±14.001比24.000±2.582、16.464±0.813比12.334±0.681,t=4.789、5.422、6.745,P<0.05),而胶质纤维酸性蛋白(GFAP)标记的阳性细胞数低于NSCs组(22.002±20.331比42.330±3.575,t=5.688,P<0.05)。NEP1-40-NSCs组的轴突新增面积大于NSCs组[(9096.000±4292.000)μm2比(4803.000±541.300)μm2,t=7.929,P<0.05];鬼笔环肽染色结果显示与NSCs组比较,NEP1-40-NSCs组的生长锥面积增宽[(103.000±79.670)μm2比(23.330±8.192)μm2,t=9.725,P<0.05],伪足数量增多且长度增加[(0.122±0.037)μm比(0.085±0.008)μm,t=4.513,P<0.05]。结论NEP1-40-NSCs在体外能够促进神经元轴突的延长,影响生长锥的结构发育,具有促进脊髓损伤修复的潜力。Objective To investigate the effect of NogoA extracellular peptide residues 1-40(NEP1-40)modified neural stem cells(NEP1-40-NSCs)on promoting neuronal axonal regeneration.Methods NEP1-40-NSCs were constructed and the levels of NEP1-40 were detected,and the differentiation potential of NEP1-40-NSCS was identified.NEP1-40-NSCs group and NSCs group were set up,and the new axon area of neurons in each group was measured after co-culture with rat hippocampal neurons.The growth cone and filopodia structures were observed by Phalloidine staining.The results were compared between two groups by t test,and compared between multiple groups by one-way ANOVA.Results The number of β3-Tubulin(Tuj-1),oliendrocyte transcription factor 2(Oligo2)and myelin basic protein(MBP)labeled positive cells in the NEP1-40-NSCs group was greater than that in the control group(47.001±16.670 vs.30.333±3.480,38.001±14.001 vs.24.000±2.582,16.464±0.813 vs.12.334±0.681,t=4.789,5.422,6.745,P<0.05),while the number of glial fibrillary acidic protein(GFAP)labeled positive cells was less than that in the control group(22.002±20.331 vs.42.330±3.575,t=5.688,P<0.05).The new axon area in the NEP1-40-NSCs group was larger than that in the NSCs group[(9096.000±4292.000)μm2 vs.(4803.000±541.300)μm2,t=7.929,P<0.05).As compared with the NSCs group,the growth cone area was widened[(103.000±79.670)μm2 vs.(23.330±8.192)μm2,t=9.725,P<0.05],the number and length of pseudopodia were increased in the NEP1-40-NSCs group[(0.122±0.037)μm vs.(0.085±0.008)μm,t=4.513,P<0.05].Conclusion NEP1-40-NSCs can promote axon elongation and affect the structural development of growth cones in vitro.

关 键 词：脊髓损伤 勿动蛋白A胞外肽残基1-40 神经干细胞 基因工程 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]