人胚胎脑海马区神经前体细胞移植到损伤的成年大鼠脑内存活及迁移的研究

THE STUDY OF THE SURVIVAL AND MIGRATION OF THE NEURAL PRECURSOR CELLS DERIVED FROM HUMAN EMBRYONIC HIPPOCAMPUS AFTER TRANSPLANTATION INTO INJURED ADULT RAT BRAIN

本文从人胚胎脑海马区分离、培养、鉴定了神经前体细胞（neural precursor cells，NPCs），并初步观察了大鼠纹状体海人酸（kainic acid，KA）损伤后，人神经前体细胞（hNPCs）移植到成年大鼠侧脑室和纹状体后存活和迁移的情况。取胎龄8～12周的人胚胎脑海马区细胞，用含人表皮生长因子（human epidermal growth factor，h-EGF）、人碱性成纤维细胞生长因子（human basic fibroblast growth factor，h-bFGF）以及人白细胞抑制因子（human leukemia inhibitory growth factor，h-LIF）的DMEM／F12培养液离体培养，以含1％胎牛血清（fetal bovine serum，FBS）的DMEM／F12诱导分化，巢蛋白（nestin）免疫荧光染色鉴定NPCs的特征。向成年大鼠右侧纹状体内立体定位注射KA，造成大鼠纹状体局部损伤。用溴脱氧尿嘧啶核苷（bromodeoxyuridine，BrdU）体外标记第2代hNPCs，48h后分别移植到正常成年大鼠和损伤大鼠手术侧的侧脑室和纹状体。6周后用抗BrdU抗体检测HNPCs的存活和迁移。结果显示：体外培养呈悬浮球状生长的细胞是nestin阳性的hNPCs。hNPCs移植6周后，在大鼠损伤侧的纹状体和侧脑室内，均检测到了BrdU阳性细胞，并有一部分细胞迁移到了周围纹状体实质和胼胝体。结果提示：体外分离培养的hNPCs移植到成年大鼠脑损伤区周围可以存活和迁移，损伤区域可能对移植细胞的迁移有一定的诱向作用。

We isolated, cultured and identified neural precursor cells （NPCs） from human embryonic hippocampus, and investigated that the survival and migration of human neural precursor cells （hNPCs） which were transplanted into unilateral lateral ventricle and striatum of KA injured rat brain. The ceils from the hippocampus of abortion human embryo （8 - 12 weeks of post-conception） were mechanically dissociated and cultured in DMEM/F12 with human epidermal growth factor （h-EGF） , human basic fibroblast growth factor （h-bFGF） and human leukemia inhibitory growth factor （h-LIF） in vitro. After inducing differentiation by DMEM/F12 containing 1% FBS, we examined the characteristics of the NPCs with nestin immunofluorescence staining. The local striatum injury was produced by unilateral stereotaxic microinjection of KA into the striatum. In vitro the passage 2 hNPCs were labeled by bromodeoxyuridine （BrdU） , after 48 h the labeled hNPCs were transplanted into the lateral ventricle and striatum of normal and injured rat brain respectively. After survival of 6 weeks, the survival and migration of transplanted cells were identified by anti-BrdU antibody. The results revealed that the floated cell spheres cultured in vitro were nestin-positive stained hNPCs. After transplantation of 6 weeks, BrdU-positive cells were found in lateral ventricle and striaturn of normal and injured rat brain. And a part of BrdU-positive cells migrated into the adjacent striatum parenchyma and corpus callosum. The study indicates that the hNPCs isolated and cultured in vitro could survive and migrate after transplanted into lateral ventricle and striaturn of rat brain with local KA injury of striatum. The lesion site in the rat brain may offer an inducement to the migration of the hNPCs.