Karbanová Jana, Mokrý Jaroslav, Kotingová Lenka
Department of Histology and Embryology, Charles University in Prague, Czech Republic.
Biomed Pap Med Fac Univ Palacky Olomouc Czech Repub. 2004 Dec;148(2):217-20.
Neural stem cells (NSCs) are tissue-specific stem cells with self-renewal potential that can give rise to neurons and glia in vivo and in vitro. The aim of this study was to transplant NSCs as whole neurospheres into intact brain and assess the fate and phenotype of their progeny generated in vivo. We isolated NSCs from E14 foetal rat forebrains and cultured them in basic fibroblast and epidermal growth factor-supplemented serum-free medium in the form of neurospheres in vitro. Neurospheres were transplanted into the intact brains of 2 Wistar rats and after a period of 3 weeks, grafted brains were examined immunohistochemically. Neurospheres formed solid grafts that were found in the lateral ventricle and in the velum interpositum under the hippocampus. The majority of cells in the transplanted tissue were identified as beta-III-tubulin(+), NeuN(+), PanNF(+) and synaptophysin(+) neurons and were accumulated throughout the graft centre. GFAP(+) astrocytes were scattered throughout the entire graft and astrocyte processes delimited the outer and perivascular surfaces. A great number of NG2(+) oligodendrocyte precursors was detected. Nestin(+) endothelial cells were found to line capillaries growing in the transplant. These data indicate that nestin(+) NSCs prevailing in neurospheres differentiate following transplantation into nestin(-) neuronal and glial cells which confirms the multipotency of NSCs. Three weeks posttransplantation neuronal and astrocyte cells reached terminal differentiation (formation of synaptic vesicles and superficial and perivascular limiting membranes) while elements of oligodendroglial cell lineage remained immature. Grafting stem cells as non-dissociated neurospheres provide cells with favourable conditions which facilitate cell survival, proliferation and differentiation. However, in the intact brain, grafted neurosphere cells were not found to integrate with the brain parenchyma and formed a compact structure demarcated from its surroundings.
神经干细胞（NSCs）是具有自我更新潜能的组织特异性干细胞，在体内和体外均可分化产生神经元和神经胶质细胞。本研究的目的是将神经干细胞作为完整的神经球移植到完整大脑中，并评估其在体内产生的子代细胞的命运和表型。我们从E14胎鼠前脑中分离出神经干细胞，并在添加碱性成纤维细胞生长因子和表皮生长因子的无血清培养基中，于体外将其培养成神经球。将神经球移植到2只Wistar大鼠的完整大脑中，3周后，对移植后的大脑进行免疫组织化学检查。神经球形成了实性移植物，位于侧脑室和海马下方的中间帆中。移植组织中的大多数细胞被鉴定为β-III-微管蛋白（+）、神经元核抗原（NeuN）（+）、泛神经丝蛋白（PanNF）（+）和突触素（+）神经元，并聚集在整个移植物中心。胶质纤维酸性蛋白（GFAP）（+）星形胶质细胞散在分布于整个移植物中，星形胶质细胞的突起界定了移植物的外表面和血管周围表面。检测到大量神经胶质细胞2（NG2）（+）少突胶质细胞前体。发现巢蛋白（Nestin）（+）内皮细胞排列在移植组织中生长的毛细血管内壁。这些数据表明，神经球中占优势的巢蛋白（+）神经干细胞在移植后分化为巢蛋白（-）神经元和神经胶质细胞，这证实了神经干细胞的多能性。移植后3周，神经元和星形胶质细胞达到终末分化（形成突触小泡以及表面和血管周围的限制膜），而少突胶质细胞系的成分仍不成熟。将干细胞作为未解离的神经球进行移植，可为细胞提供有利条件，促进细胞存活、增殖和分化。然而，在完整大脑中，未发现移植的神经球细胞与脑实质整合，而是形成了一个与周围环境分界的紧密结构。
