人脐血单核细胞颈内动脉输注对血管性痴呆大鼠学习记忆的作用及在脑内的迁移分化

Effect of intracarotid administration of human cord blood mononuclear cells on learning-memory abilities in the vascular dementia rats brain and its migration, differentiation

目的观察颈内动脉输注人脐血单个核细胞（HCMNCs）透过血脑屏障在血管性痴呆（VD）大鼠脑内仔活、迁移分化及对学习记忆的影响。方法改良Pulsinellis四血管阻断法建立VD大鼠模型；体外分离HCMNCs，术后24h治疗组颈内动脉输注数量为3×10^6／0．5ml的Brdu标记细胞；采用免疫组化法和穿梭箱系统检测HCMNCs在VD大鼠脑内的存活、迁移和分化情况以及对认知功能的影响。结果HCMNCs广泛分布于VD大鼠脑组织，可存活较长时间（8周以上），随着时间的延长，迁移方向集中，主要聚集迁移至海马、大脑皮质等区域；HCMNCs在VD大鼠脑内微环境作用下2周已开始向神经细胞转化，表达人神经丝蛋白（NF）、神经元特异性烯醇化酶（NSE）以及胶质原纤维酸性蛋白（GFAP）。模型组大鼠AAR比率[（55．4±4．5），（42．1±4．5），（44．2±3．6）]低于对照组[（91．7±3．9），（90．0±4．3），（92．5±5．0）]，差异非常显著（P〈0．01）；治疗组[（67．1±3．3），（69．2±4．7），（70．8±4．7）]较模型组明显提高（P〈0．01）。结论颈内动脉输注HCMNCs可透过VD大鼠血脑屏障向脑缺血易损伤部位迁移聚集、存活，定向分化为神经样细胞，并可显著改善VD大鼠学习记忆能力。

Objective To observe the characteristics of the survival, migration,differentiation and effect of the intracarotidly infused human cord blood mononuclear cells（HCMNCs） on learning-memory abilities across the blood-brain barrier（BBB） in the vascular dementia （VD） rats. Methods The VD rat model was established by modified pulsinellis 4-vessel occlusion（4-VO）. HCMNCs were isolated from human umbilical cord blood in vitro. The treatment group received intracarotid infusion 3 × 10^6/0.5 ml HCMNCs which were prelabeled with BrdU in vitro. Immunohistochemical method and computerized shuttle-training case were used to examine their survival, migration,differentiation,and the cognitive function in the VD rats. Results HCMNCs transplanted by intracarotid administration had been found throughout the brain tissue and could survive for a long time（ exceeding 8 weeks ）. They localized to migratory direction, and the vast majority distributed in the ischemic zone of injury such as hippocampus and cerebral cortex. Immunohistochemical staining showed implanted cells began to differentiate into nerve cells expressing human neurofilament （NF） ,neuron-specific enolase （NSE） and glial fibrillary acid protein （GFAP） in the brain tissues microenvironment at 2 weeks after infusion. AAR in the model group （ 55.4 ± 4.5, 42.1 ± 4.5,44.2 ± 3.6） showed significantly decrease compared with the control （ 91.7 ± 3.9,90.0 ± 4.3,92.5 ± 5.0, P 〈 0.01 ）, and AAR in the treatment （ 67.1 ± 3.3,69.2 ± 4.7,70.8 ± 4.7 ） was significant higher than the model（ P〈0.01 ）. Conclusion Intracarotid administration of HCMNCs could enter the VD rats cerebra parenchyma across BBB, and migrate into the damaged brain tissue to localize, survive and differentiate into nerve cells,and significantly improve learning-memory abilities of VD rats.