中国组织工程研究与临床康复
中國組織工程研究與臨床康複
중국조직공정연구여림상강복
JOURNAL OF CLINICAL REHABILITATIVE TISSUE ENGINEERING RESEARCH
2010年
6期
985-991
,共7页
黄作义%吴成吉%朱晓峰%董淑欣%魏春杰
黃作義%吳成吉%硃曉峰%董淑訢%魏春傑
황작의%오성길%주효봉%동숙흔%위춘걸
电刺激%小脑顶核%神经干细胞%移植%存活%迁移
電刺激%小腦頂覈%神經榦細胞%移植%存活%遷移
전자격%소뇌정핵%신경간세포%이식%존활%천이
背景:缺血早期半暗带中神经元和内皮细胞的坏死和凋亡无法得到缓解,加之移植后的存活率低,向功能细胞的分化率低,是单纯神经干细胞移植的缺陷.课题组提出整体干预理念,期望给移植细胞提供优化的整体环境.目的:观察神经干细胞移植与电刺激小脑顶核相结合整体干预对移植神经干细胞存活与迁移的影响.方法:体外分离、培养新生Wistar大鼠海马表皮生长因子反应性神经干细胞,Brdu标记,并用胎牛血清诱导,观察其多向分化潜能;80只Wistar大鼠分为4组:顶核刺激+神经干细胞移植组(n=32):左侧小脑顶核刺激24 h后行右侧大脑中动脉梗死,再24 h后将神经干细胞立体定向注入右侧侧脑室内;顶核刺激组(n=8):PBS代替神经干细胞,其余同顶核刺激+神经干细胞移植组;单纯神经干细胞移植组(n=32):同心圆电极插入小脑顶核,不通电其余同顶核刺激+神经干细胞移植组;对照组(n=8):同心圆电极插入小脑顶核后不通电,其余同顶核刺激组.记录梗死后6,24 h,3,7,14,28 d大鼠的神经功能;分别在移植后3,7,14,28 d处死大鼠,以梗死灶为中心切片,Brdu免疫组织化学染色,观察移植神经干细胞的存活与迁移.结果与结论:分离培养的表皮生长因子反应性细胞表达nestin抗原,并有自我更新及多向分化潜能,在胎牛血清诱导下可分化为神经胶质细胞和神经元.经Brdu标记后,85%以上神经干细胞表达Brdu抗原.移植28 d内顶核刺激+神经干细胞移植组功能评分明显优于与其他3组(P < 0.05～0.01).顶核刺激+神经干细胞移植组在移植14,28 d存活细胞数明显多于单纯神经干细胞移植组;提示电刺激小脑顶核可显著提高移植神经干细胞的存活率及大脑中动脉梗死大鼠的神经功能评分;优化的整体环境可提高移植神经干细胞对局灶性脑缺血损伤细胞的替代作用.
揹景:缺血早期半暗帶中神經元和內皮細胞的壞死和凋亡無法得到緩解,加之移植後的存活率低,嚮功能細胞的分化率低,是單純神經榦細胞移植的缺陷.課題組提齣整體榦預理唸,期望給移植細胞提供優化的整體環境.目的:觀察神經榦細胞移植與電刺激小腦頂覈相結閤整體榦預對移植神經榦細胞存活與遷移的影響.方法:體外分離、培養新生Wistar大鼠海馬錶皮生長因子反應性神經榦細胞,Brdu標記,併用胎牛血清誘導,觀察其多嚮分化潛能;80隻Wistar大鼠分為4組:頂覈刺激+神經榦細胞移植組(n=32):左側小腦頂覈刺激24 h後行右側大腦中動脈梗死,再24 h後將神經榦細胞立體定嚮註入右側側腦室內;頂覈刺激組(n=8):PBS代替神經榦細胞,其餘同頂覈刺激+神經榦細胞移植組;單純神經榦細胞移植組(n=32):同心圓電極插入小腦頂覈,不通電其餘同頂覈刺激+神經榦細胞移植組;對照組(n=8):同心圓電極插入小腦頂覈後不通電,其餘同頂覈刺激組.記錄梗死後6,24 h,3,7,14,28 d大鼠的神經功能;分彆在移植後3,7,14,28 d處死大鼠,以梗死竈為中心切片,Brdu免疫組織化學染色,觀察移植神經榦細胞的存活與遷移.結果與結論:分離培養的錶皮生長因子反應性細胞錶達nestin抗原,併有自我更新及多嚮分化潛能,在胎牛血清誘導下可分化為神經膠質細胞和神經元.經Brdu標記後,85%以上神經榦細胞錶達Brdu抗原.移植28 d內頂覈刺激+神經榦細胞移植組功能評分明顯優于與其他3組(P < 0.05～0.01).頂覈刺激+神經榦細胞移植組在移植14,28 d存活細胞數明顯多于單純神經榦細胞移植組;提示電刺激小腦頂覈可顯著提高移植神經榦細胞的存活率及大腦中動脈梗死大鼠的神經功能評分;優化的整體環境可提高移植神經榦細胞對跼竈性腦缺血損傷細胞的替代作用.
배경:결혈조기반암대중신경원화내피세포적배사화조망무법득도완해,가지이식후적존활솔저,향공능세포적분화솔저,시단순신경간세포이식적결함.과제조제출정체간예이념,기망급이식세포제공우화적정체배경.목적:관찰신경간세포이식여전자격소뇌정핵상결합정체간예대이식신경간세포존활여천이적영향.방법:체외분리、배양신생Wistar대서해마표피생장인자반응성신경간세포,Brdu표기,병용태우혈청유도,관찰기다향분화잠능;80지Wistar대서분위4조:정핵자격+신경간세포이식조(n=32):좌측소뇌정핵자격24 h후행우측대뇌중동맥경사,재24 h후장신경간세포입체정향주입우측측뇌실내;정핵자격조(n=8):PBS대체신경간세포,기여동정핵자격+신경간세포이식조;단순신경간세포이식조(n=32):동심원전겁삽입소뇌정핵,불통전기여동정핵자격+신경간세포이식조;대조조(n=8):동심원전겁삽입소뇌정핵후불통전,기여동정핵자격조.기록경사후6,24 h,3,7,14,28 d대서적신경공능;분별재이식후3,7,14,28 d처사대서,이경사조위중심절편,Brdu면역조직화학염색,관찰이식신경간세포적존활여천이.결과여결론:분리배양적표피생장인자반응성세포표체nestin항원,병유자아경신급다향분화잠능,재태우혈청유도하가분화위신경효질세포화신경원.경Brdu표기후,85%이상신경간세포표체Brdu항원.이식28 d내정핵자격+신경간세포이식조공능평분명현우우여기타3조(P < 0.05～0.01).정핵자격+신경간세포이식조재이식14,28 d존활세포수명현다우단순신경간세포이식조;제시전자격소뇌정핵가현저제고이식신경간세포적존활솔급대뇌중동맥경사대서적신경공능평분;우화적정체배경가제고이식신경간세포대국조성뇌결혈손상세포적체대작용.
BACKGROUND: The discrepancy of neural stem cells (NSCs) transplantation is low cell survival rate and poor differentiation, which can not relieve the large apoptosis of neuron and endothelial cells in ischemic penumbra. This paper proposes a principle of overall intervention to provide an optimal environment for transplanted cells. OBJECTIVE: To observe the effect of cerebellar fastigial nucleus electrical stimulation (FNS) on survival and migration of NSCs. METHODS: The EGF-responsive hippocampal NSCs of neonate Wistar rats were isolated and cultured in vitro. They were labeled by Brdu and induced by embryo cattles blood serum. The multi-differentiation potential was studied. Totally 80 rats were assigned into 4 groups. FNS+NSCs transplantation (n=32): at 24 hours FNS, right middle cerebral artery occlusion (MCAO) was prepared and received NSCs transplantation. FNS group (n =8): received the same procedure as FNS+NSCs transplantation except substitute PBS for NSCs. NSCs transplantation group (n=32): concentric electrode was inserted without electrify, the remained procedure was the same to FNS+NSCs transplantation group. Control group (n=8): concentric electrode was inserted without electrify, the remained procedure was the same to FNS group. The neural functions were recorded at hours 6, 24 and days 3, 7, 14, 28 after infarction. Rats were sacrificed at days 3, 7, 14 and 28 after transplantation, and the survival and migration of NSCs were investigated by Brdu immunocytochemical staining. RESULTS AND CONCLUSION: Isolated and purified epidermal growth factor-responsive hippocampal NSCs were Nestin-positive and had ability of self-renewing and multi-directional differentiation. More than 85% NSCs expressed the antigen of Brdu after Brdu labeled. The functional scores of the FNS+NSCs transplantation group were significant better than those of the other 3 groups at 28 days after transplantation (P < 0.05-0.01). The number of survival cells in the FNS+NSCs transplantation group was significantly greater than that of NSCs transplantation group at days 14 and 28 after transplantation. FNS can improve the survival rate of transplanted NSCs and the functional scores following MCAO. The grading-up global environment can improve the substitution role of transplanted NSCs at local cerebral infarction damage.
