大面积脑梗塞患者自体神经干细胞的培养与鉴定  

Culture differentiation and identification of autologous neural stem cells in patients with massive cerebral infarction

作　　者：王佳[1] 史铁钧 赵全军 王涛[1] 崔绍杰 任飞 杨凯 Wang Jia;Shi Tiejun;Zhao Quanjun;Wang Tao;Cui Shaojie;Ren Fei;Yang Kai(The Stereotactic and Neurofunctional Center of 306^th Clinical College of PLA, Anhui Medical University;Department of Teaching, The 306th Hospital of PLA, Beijing 100101 , China)

机构地区：[1]解放军第三〇六医院安徽医科大学解放军第三〇六临床学院立体定向及脑功能性疾病诊治中心 [2]解放军第三〇六医院科训科,北京100101

出　　处：《神经解剖学杂志》2019年第3期300-304,共5页Chinese Journal of Neuroanatomy

基　　金：首都临床特色应用研究与成果推广(Z161100000516199)

摘　　要：目的:通过改进原代培养方法,从大面积脑梗塞患者废弃脑组织中培养、传代和鉴定自体神经干细胞(NSCs),并进行诱导分化研究。方法:收集2例右侧大面积脑梗塞患者手术中废弃脑组织各1 g以上,按改良方法进行原代培养并传代,通过细胞免疫荧光染色检测NSCs标志物巢蛋白(nestin)的表达。传代培养至7代时选取部分细胞进行诱导分化研究,通过细胞免疫荧光染色检测胶质纤维酸性蛋白(GFAP)和微管蛋白(β-tubulin)的表达来评估NSCs分化情况。结果:培养3~7 d后,2例患者废弃脑组织中均可见干细胞球生长,nestin检测呈阳性表达。经传代培养至第10代后,仍可维持其神经干细胞特征,选取7代贴壁细胞经β-Tubulin及GFAP免疫荧光检测可见阳性表达。结论:成功从大面积脑梗塞患者前颞叶(邻近于海马区)废弃脑组织中分离培养获得自体NSCs,并可向神经元样细胞及星形胶质样细胞分化。Objective: Culture,identification and differentiation of autologous neural stem cells(NSCs) from the abandoned brain tissues in patients of massive cerebral infarction by the improvement of primary culture. Methods: The abandoned brain tissues were collected from the anterior temporal lobe in 2 cases of right massive cerebral infarction during surgery. Above 1 g abandoned brain tissues in each case were primary cultured and passaged by the improvement methods. The expression of nestin(the marker of NSCs) was detected by immunofluorescence. The induced differentiation began from the 7th passage. Expression of glial fibrillary acidic protein(GFAP) and tubulin(β-tubulin) were detected by immunofluorescence. Results: About 3 to 7 days after inoculation,the discarded brain tissues of the 2 cases were observed of nestin positive cells outgrowth. After subculture,the 10 thpassage of cells still preserved their characteristic of NSCs. The 7 thpassage of cells were collected for immunofluorescence cytochemistry in which some adherent cells were β-Tubulin and GFAP positive. Conclusion: Adult neural stem cells are enriched in the subventricular zone and hippocampus. This study successfully obtained autologous NSCs from the abandoned brain tissues of the anterior temporal lobe(near the hippocampus) in patients of massive cerebral infarction and the autologous NSCs could be differentiated to neuron-like cells and astrocyte-like cells.

关 键 词：神经干细胞 细胞培养 脑梗塞 脑卒中 

分 类 号：R743.3[医药卫生—神经病学与精神病学]