CAJ | 학술논문

背景:以往对脑缺血再灌注后半胱氨酸天冬氨酸蛋白酶3(cysteinyl aspartate specific protease 3,Caspase-3)的表达均缺乏长时间的动态观察.目的:探讨胶质细胞源性神经营养因子基因修饰的神经干细胞移植对暂时性脑缺血大鼠脑组织Caspase-3表达的影响.设计:随机对照动物实验.材料:清洁级成年SD大鼠60只,随机分为4组:正常对照组5只、缺血再灌注组5只、单纯细胞移植组25只、基因修饰细胞移植组25只.另取清洁级新生SD大鼠数只,用于分离培养神经干细胞.方法:除正常对照组外,其余3组大鼠按改良性线栓法制备暂时性脑缺血模型.再灌注3 d,单纯细胞移植组从右侧侧脑室每只注入20 μL神经干细胞悬液,含(4.0-5.0)×10~5个神经干细胞;基因修饰细胞移植组每只注入等量胶质细胞源性神经营养因子基因修饰的神经干细胞悬液;缺血再灌注组每只注入20μL生理盐水.正常对照组、缺血再灌注组在缺血再灌注1周麻醉处死动物,单纯细胞移植组、基因修饰细胞移植组分别在缺血再灌注1,2,3,5,7周麻醉处死动物,5只/时间点.主要观察指标:采用免疫组织化学SP法观察Caspase-3在海马和额顶皮质表达及分布特点.结果:免疫组织化学染色显示,Caspase-3免疫阳性产物位于细胞核、细胞质和部分突起.在海马:单纯细胞移植组、基因修饰细胞移植组Caspase-3阳性细胞数均随再灌注时间的延长而逐渐减少;除缺血再灌注第1周外,其余各时间点基因修饰细胞移植组Caspase-3阳性细胞数均明显少于单纯细胞移植组(P<0.05).在额顶皮质:单纯细胞移植组、基因修饰细胞移植组Caspase-3阳性细胞数亦均随再灌注时间的延长而逐渐减少;除缺血再灌注第1,2周外,其余各时间点基因修饰细胞移植组Caspase-3阳性细胞数均明显少于单纯细胞移植组(P<0.05).结论:胶质细胞源性神经营养因子基因修饰的神经干细胞移植通过降低Caspase-3的表达,减少神经元凋亡,从而抑制缺血性脑损伤的进行性恶化,其移植效果优于单纯神经干细胞移植. 揹景:以往對腦缺血再灌註後半胱氨痠天鼕氨痠蛋白酶3(cysteinyl aspartate specific protease 3,Caspase-3)的錶達均缺乏長時間的動態觀察.目的:探討膠質細胞源性神經營養因子基因脩飾的神經榦細胞移植對暫時性腦缺血大鼠腦組織Caspase-3錶達的影響.設計:隨機對照動物實驗.材料:清潔級成年SD大鼠60隻,隨機分為4組:正常對照組5隻、缺血再灌註組5隻、單純細胞移植組25隻、基因脩飾細胞移植組25隻.另取清潔級新生SD大鼠數隻,用于分離培養神經榦細胞.方法:除正常對照組外,其餘3組大鼠按改良性線栓法製備暫時性腦缺血模型.再灌註3 d,單純細胞移植組從右側側腦室每隻註入20 μL神經榦細胞懸液,含(4.0-5.0)×10~5箇神經榦細胞;基因脩飾細胞移植組每隻註入等量膠質細胞源性神經營養因子基因脩飾的神經榦細胞懸液;缺血再灌註組每隻註入20μL生理鹽水.正常對照組、缺血再灌註組在缺血再灌註1週痳醉處死動物,單純細胞移植組、基因脩飾細胞移植組分彆在缺血再灌註1,2,3,5,7週痳醉處死動物,5隻/時間點.主要觀察指標:採用免疫組織化學SP法觀察Caspase-3在海馬和額頂皮質錶達及分佈特點.結果:免疫組織化學染色顯示,Caspase-3免疫暘性產物位于細胞覈、細胞質和部分突起.在海馬:單純細胞移植組、基因脩飾細胞移植組Caspase-3暘性細胞數均隨再灌註時間的延長而逐漸減少;除缺血再灌註第1週外,其餘各時間點基因脩飾細胞移植組Caspase-3暘性細胞數均明顯少于單純細胞移植組(P<0.05).在額頂皮質:單純細胞移植組、基因脩飾細胞移植組Caspase-3暘性細胞數亦均隨再灌註時間的延長而逐漸減少;除缺血再灌註第1,2週外,其餘各時間點基因脩飾細胞移植組Caspase-3暘性細胞數均明顯少于單純細胞移植組(P<0.05).結論:膠質細胞源性神經營養因子基因脩飾的神經榦細胞移植通過降低Caspase-3的錶達,減少神經元凋亡,從而抑製缺血性腦損傷的進行性噁化,其移植效果優于單純神經榦細胞移植.
배경:이왕대뇌결혈재관주후반광안산천동안산단백매3(cysteinyl aspartate specific protease 3,Caspase-3)적표체균결핍장시간적동태관찰.목적:탐토효질세포원성신경영양인자기인수식적신경간세포이식대잠시성뇌결혈대서뇌조직Caspase-3표체적영향.설계:수궤대조동물실험.재료:청길급성년SD대서60지,수궤분위4조:정상대조조5지、결혈재관주조5지、단순세포이식조25지、기인수식세포이식조25지.령취청길급신생SD대서수지,용우분리배양신경간세포.방법:제정상대조조외,기여3조대서안개량성선전법제비잠시성뇌결혈모형.재관주3 d,단순세포이식조종우측측뇌실매지주입20 μL신경간세포현액,함(4.0-5.0)×10~5개신경간세포;기인수식세포이식조매지주입등량효질세포원성신경영양인자기인수식적신경간세포현액;결혈재관주조매지주입20μL생리염수.정상대조조、결혈재관주조재결혈재관주1주마취처사동물,단순세포이식조、기인수식세포이식조분별재결혈재관주1,2,3,5,7주마취처사동물,5지/시간점.주요관찰지표:채용면역조직화학SP법관찰Caspase-3재해마화액정피질표체급분포특점.결과:면역조직화학염색현시,Caspase-3면역양성산물위우세포핵、세포질화부분돌기.재해마:단순세포이식조、기인수식세포이식조Caspase-3양성세포수균수재관주시간적연장이축점감소;제결혈재관주제1주외,기여각시간점기인수식세포이식조Caspase-3양성세포수균명현소우단순세포이식조(P<0.05).재액정피질:단순세포이식조、기인수식세포이식조Caspase-3양성세포수역균수재관주시간적연장이축점감소;제결혈재관주제1,2주외,기여각시간점기인수식세포이식조Caspase-3양성세포수균명현소우단순세포이식조(P<0.05).결론:효질세포원성신경영양인자기인수식적신경간세포이식통과강저Caspase-3적표체,감소신경원조망,종이억제결혈성뇌손상적진행성악화,기이식효과우우단순신경간세포이식.
BACKGROUND: There is few studies addressing the long-playing dynamic observation of cysteinyl aspartate specific protease 3 (Caspase-3) expression following cerebral ischemia/reperfusion.OBJECTIVE: To investigate the effect of transplantation of the neural stem cells (NSCs) modified with gene of glial cell line-derived neurotrophic factor (GDNF) on expression of Caspase-3 in adult Sprague Dawley rats with transient cerebral ischemia.DESING: Randomized controlled animal study.MATERIALS: Sixty Sprague Dawley rats were divided randomly into normal control group (N, n =5), ischemia/reperfusion group (IR, n=5), neural stem cell group (NSCs, n=25) and NSCs modified with gene of GDNF group (GDNF/NSCs, n =25). Several clean neonatal Sprague-Dawley rats were selected to harvest NSCs.METHODS: With the exception of normal control group, models of transient cerebral ischemia were created by modified suture method in other groups. At day 3 following reperfusion, 20 μL NSC suspension containing (4.0-5.0)×10~5 NSCs was infused into rats of the NSC group via right lateral ventricle. An equal volume of GDNF-modified NSC suspension was injected into rats of the GDNF/NSC group. 20 μL saline was infused into the rats of the ischemia/reperfusion group. Animals were anesthetized and sacrificed at week 1 following ischemia/reperfusion in the normal control and ischemia/reperfusion groups. Animals were anesthetized and sacrificed at weeks 1, 2, 3, 5, 7 following ischemia/reperfusion in the NSC and GDNF/NSC groups, 5 rats in each time point.MAIN OUTCOME MEASURES: The strept avidin-biotin immunostaining method was used to observe the distributive characteristics of Caspase-3 in the hippocampus and frontal parietal cortex.RESULTS: Immunohistochemical method (SP) showed that positive capase-3 products expressed in nucleus, cytoplasm and partial neurite. In hippocampus, number of Caspase-3-positive cells was decreased in NSC and GDNF/NSC groups. With the exception of at 1-week reperfusion, number of Caspase-3-positive cells was significantly lessened in the GDNF/NSC group compared with the NSC group at other time points (P < 0.05). In frontoparietal cortex, number of Caspase-3-positive cells was reduced in the NSC and GDNF/NSC groups over time. Except 1 and 2 weeks following ischemia/reperfusion, number of Caspase-3-positive cells was significantly lessened in the GDNF/NSC group compared with the NSC group (P < 0.05).CONCLUSION: Transplanting NSCs modified with gene of GDNF can improve remarkably neural function by deceasing Caspase-3 expression and reducing the nervous cell apoptosis. The transplantation of NSCs modified with gene of GDNF obtained better outcomes compared with NSC transplantation.