成体小鼠脊髓实质神经干细胞的分离培养及其多能性的鉴定

目的从成年小鼠脊髓实质中分离培养并鉴定成体神经干细胞。方法采用无血清培养基及单细胞克隆技术，从成年小鼠脊髓实质中分离培养获得大量具备单细胞克隆能力的细胞，经免疫荧光染色证实单克隆细胞表达Nestin抗原并在分化后表达各种特异性的神经细胞抗原。结果从成年小鼠脊髓实质中分离培养获得的具备连续克隆能力的细胞可表达神经干细胞的Nestin抗原，并且分化后表达神经元及胶质细胞的特异性抗原。结论成功地从成年小鼠的脊髓实质中分离出具备增殖分化能力的成体神经干细胞，为进一步利用其进行脊髓损伤治疗的研究提供了实验基础。

Isolation , incubation and identification of parenchymal neural stem cells in adult mouse spinal cord

Objective To isolate and identify the adult neural stem cells from the parenchyma of spinal cord in adult mouse. Methods The parenchymal spinal cord from adult mouse was dissected and dissociated by mechanical trituration. The tissue suspension was cultured in serum-free DMEM/F12 medium supplemented with EGF and B27. The cell colonies generated from a single cell were screened by limited dilution and incubated with BrdU. The cell colonies were transferred into medium with serum to induce differentiation. The cells were identified with antibodies to Nestin, BrdU, MAP2 and GFAP by immunofluorescence staining. Results The cells were cultured for seven clays to generate proliferative neurospheres. The majority of cells in these neurospheres expressed Nestin and were differentiated into MAP2-positive cells and GFAP-positive cells in medium containing with fetal bovine serum. Conclusion A significant number of neural stem cells are present in the paxenchymal adult mouse spinal cord and can proliferate and also give rise to neurons and glia in vitro.