神经祖细胞与纤维蛋白支撑物移植对老年痴呆模型鼠的记忆认知功能改善及其机理探究

目的探讨阿尔茨海默病（AD）模型鼠双侧海马区移植含多因子孵育的神经祖细胞（NPCs）后记忆认知功能改善情况及NPCs移植后迁移定位和分化能力。 方法取胎龄10?d的C57BL/6J孕鼠，分离得胎鼠NPCs，NPCs体外分化及鉴定，AD模型鼠分3组：NPCs+因子组、因子组及PBS组，对照组为同月龄C57BL/6J小鼠；Morris水迷宫及新物体识别实验检测AD模型鼠移植NPCs细胞前，及移植1至6个月后记忆行为变化情况；通过免疫荧光，免疫组化和Western-blot检测海马区移植的NPCs向神经元和胆碱能神经元分化及迁移能力。组间比较采用F检验。 结果Morris水迷宫实验中，NPCs+因子组找到平台前的逃避潜伏期时间（14.12±7.45）s要明显低于注射PBS的AD模型鼠组[（39.65±4.64）?s，F = 2.578，P = 0.0094]，因子组时间（15.68±5.34）s同样低于PBS组[（39.65±4.64）s，F?= 1.324，P = 0.0016]，24 h撤去平台后，NPCs+因子组逃避潜伏期时间（15.12±3.52）s仍低于PBS组[（37.17±2.18）?s，F = 2.598，P = 0.0003]，因子组时间（16.62±3.23）s同样低于PBS组[（37.17±2.18）s，F = 2.186，P = 0.0004）]；新物体识别实验中，各实验组对新物体探究时间占总探究时间百分比结果中，NPCs+因子组（68.46±2.4）%要高于PBS组[（54.47±4.79）%，F =3.983，P = 0.018]，因子组（65.20±1.03）%同样高于PBS组[（54.47±4.79）%，F = 21.63，P = 0.042]；实验结果表明，通过移植细胞与因子AD模型鼠的记忆认知功能在早期均得到改善，随着时间的增长，移植NPCs组的记忆改善情况持续时间更长久；Western blot结果显示AD模型鼠海马区胆碱能神经元与正常C57BL/6J鼠相比表达减少，移植NPCs后，AD模型鼠脑内胆碱能神经元增多；免疫荧光与免疫组化结果显示，移植的NPCs在AD模型鼠脑内移植区存活，并向胆碱能神经元分化。 结论AD模型鼠双侧海马区移植的含多因子孵育的NPCs，通过分化成功能性的胆碱能神经元来改善AD鼠的记忆认知功能。

Study on the memory cognitive function of neural progenitor cell transplantation with fibrin suppository in Alzheimer's disease model rats and its mechanism

ObjectiveTo investigate the improvement of memory function and the differentiation ability after transplantation of NPCs with multiple factor to hippocampus in AD model rats. MethodFetal rat NPCs were isolated from C57BL/6J pregnant rats at gestational age of 10 days; AD model mice were divided into three groups: NPCs+ multiple factors group, multiple factors group and PBS group, and The control group is the same age C57BL/6J mouse. Morris water maze (MWM) and novel object recognition task were used to detect the changes of memory behavior before and after transplanting NPC cells in AD model mice; Immunofluorescence, immunohistochemistry and Western-blot were used to detect the differentiation and migration ability of NPCs transplanted into hippocampus. GraphPadPrism 7.0 software was used for statistical analysis using t test. ResultIn the Morris water maze experiment, the escape latency (14.12 ± 7.45)?s before the platform of the mice injected with NPCs and factors was found to be significantly lower than that of the AD model mice injected with PBS [(39.65±4.64)?s, F?= 2.578, P = 0.0094]. After 24 hours with platform removed, the escape latency of the mice injected with NPCs and factors (15.12±3.52)?s was still lower than that of PBS group [(37.17±2.18)?s, F?= 2.598, P = 0.0003], and the time of factor group (16.62±3.23)?s was also lower than that of the PBS group[(37.17±2.18)?s, F = 2.186, P = 0.0004)]. In the new object recognition experiment, the NPC + factor group was better (68.46±2.4)% than the PBS group [(54.47±4.79)%, F = 3.983, P?= 0.018]. The factor group (65.20±1.03)% was also better than the PBS group [(54.47±4.79)%, F?= 21.63, P = 0.042]. Results show that, the memory cognitive function AD mice was improved at an early stage of treatment. Over time, the memory improvement of the NPC group lasted longer. Western blot results showed a decrease in cholinergic neurons in the hippocampus of AD model mice compared with normal C57 mice. After transplantation of NPCs, the number of cholinergic neurons in the brain of AD model increased. Immunofluorescence and immunohistochemistry showed that, the transplanted NPCs survived in the brain and differentiated into cholinergic neurons. ConclusionFactor-incubated NPCs transplanted in bilateral hippocampus of AD model rats improve memory cognitive function of AD rats by differentiating into functional cholinergic neurons.