脐血CD133^+移植改善转基因痴呆小鼠认知和存活能力(英文)

Cord blood CD133^+ cell transplantation improves cognitive function and survival of transgenic mice with dementia

背景:脐血中富含早期干细胞的一个亚群CD133+细胞,该亚群是具有广泛的增生和分化潜能的原始细胞,被认为具有向神经细胞分化的潜能。目的:假设脐血CD133+细胞对改善痴呆小鼠的认知功能和存活有使用价值,检测脐血CD133+细胞移植后痴呆鼠的认知和存活功能的变化,拟予验证。设计、时间和单位:完全随机区组设计的动物实验,于2005-09/2007-01在天津血液学研究所完成。材料:选用48只雄性APP695转基因小鼠,随机分为3组:对照组(n=8)、CD133+细胞移植组(n=20)和CD133-细胞移植组(n=20)。方法:对照组小鼠经脑室注射10μL磷酸缓冲液(PBS),CD133+细胞移植组和CD133-细胞移植组分别经脑室注射10μLCD133+(5×104/μL)和CD133-(5×104/μL)脐血细胞移植。主要观察指标:以水迷宫实验评价转基因小鼠在细胞移植后的认知功能,并记录移植后小鼠的存活时间。以Dil荧光标记法和免疫组化检测移植细胞的分化类型。结果:在移植后30d,CD133+细胞移植组小鼠的认知功能明显好于CD133-细胞移植组和对照组,差异有显著性意义(P<0.05),这种改善效果在移植后180d仍然存在(P<0.05)。CD133+细胞移植组小鼠平均存活时间比CD133-细胞移植组和对照组明显延长,差异有显著性意义(P<0.05)。标记有Dil的脐血细胞在移植后30d已经迁移到多个脑区,其中主要在双侧顶叶皮质和海马区,并且能够表达神经细胞标志Ⅲ型β微管蛋白、神经微丝、神经烯醇化酶和胶质酸性蛋白。CD133-细胞移植组脑切片内,Dil标记的脐血源细胞主要分布在侧脑室及其周围,很少能检测到阳性胶质酸性蛋白,Ⅲ型β微管蛋白,或神经烯醇化酶的脐血细胞。移植后30d,CD133+细胞移植组Dil标记的脐血细胞表达III型β微管蛋白的百分率明显高于180d,表达神经烯醇化酶的脐血细胞百分率明显低于180d,差异均有显著性意义(P<0.01)。结论:实验结果提示,移植CD133+细胞后出现的对认知功能改善和对存活时间延长的作用,可能主要归功于移植细胞对功能不良细胞的替代或者对神经环路的改善。

BACKGROUND： Human umbilical cord blood （CB）-derived CD133^＋ cells are a minority population of primitive cells with extensive proliferation and differentiation potentials, which are considered to have ability of neural differentiation. OBJECTIVE： We hypothesized a possible application of CB CD 133^＋ cells in the cognitive and survival function of mice with dementia, the present study observed the changes of the cognitive function and survival of amyloid precursor protein（APP） transgenic mice after CB CD133^＋ ceils transplantation to verify the above assumption. DESIGN, TIME AND SETTING： A completely randomized block design of animal experiments was performed in the Hematology Institute of Tianjin Hematology Hospital from September 2005 to December 2007. MATERIALS： Forty-eight eight-month-old male APP 695 transgenic C57BL/6 （BDFI/KM） mice were selected in this experiments All mice were divided randomly into three groups： control group （n=8）, CD 133^＋ transplantation group （n=20） and CD 133^＋ transplantation group （n=20）. METHODS： Mice in control groups received an intraventricular injection of 10μL phosphate buffered saline （PBS）. The transgenic mice that received an intraventricular injection of 10μL CD133^＋ （5×10^4/μL） and CD133 CB cells （5× 10^4/μL） respectively. MAIN OUTCOME MEASURES： Radial arm water maze （RAWM） was used to evaluate cognitive function of the mice and the survival days of mice in different groups were recorded. Immunohistochemical assessments and Dil Fluorescence labeled way was used to detect the differentiation phenotype of transplanted ceils. RESULTS： The cognitive function of the mice in CD 133^＋ transplantation group was significantly improved compared with the mice in CD133^- transplantation and control groups both 30 and 180 days after transplantation （P 〈 0.05）. The mean survival time of the mice in CD133^＋ transplantation group was significantly increased compared with CD133^- transplantin group and control group （P 〈 0.05）. It was observed that the transplantation CB CD133^＋ cells labeled with Dil migrated into several brain regions at day 30 post-transplantation. These cells were stained for human β Ⅲ-tubulin, neuralfilement（NF）, neuron specific enolase （NSE）, and gfial fibriliary acidic protein（GFAP）. However, in the brain of mice that received CD133^- cells transplantation, CB cells were distributed mainly in and around the lateral ventricle at day 30 and 180 post-transplantation and GFAP-, β Ⅲ-tubulin- and NSE-positive cells were rarely detected. After intraventricular transplantation of CB CD 133^＋ cells, the percentage of transplanted Dil-labeled CB cells expressing β Ⅲ-tubulin was significant higher at day 30 than at day 180, and the percentage of CB cells expressing NSE was significant lower at day 30 than that at day 180 （both P 〈 0.015. The percentage of CB cells expressing GFAP was relatively constant between the days 30 and 180 after transplantation （P 〉 0.05）. CONCLUSION： The result of this experiment suggested that the cognitive and survival function improvement achieved by transplantation of CB CD 133^＋ cells is mainly due to a replacement of dysfunctional ceils or augmentation of neural circuit by CB CD 133^＋ cells transplantation.