人胚胎皮层神经前体细胞的分离培养及鉴定

Isolation and identification of neural precursor cells from human embryonic cortical tissue

目的 从 2 0周人胚皮层中分离神经前体细胞并鉴定其增殖及分化能力。方法 应用包含碱性成纤维细胞生长因子 (bFGF)和表皮细胞生长因子 (EGF)的无血清限制培养基从 2 0周自然流产人胚胎皮层组织中分离神经前体细胞 ,通过神经球形成法使其不断增殖 ,利用BrdU检测细胞的增殖能力 ,去除生长因子后诱导神经前体细胞分化。利用流式细胞仪检测神经球内细胞的增殖细胞周期。结果 人胚胎皮层存在神经前体细胞 ,这些细胞于EGF及bFGF作用下可不断增殖形成神经球 ,并能自我更新和结合BrdU ,于生长因子去除后能够分化成熟的神经元、星形胶质细胞和少突胶质细胞。反复传代后这些细胞可保持其增殖及分化能力。细胞周期结果显示神经球内细胞增殖较为活跃。结论 从人胚胎中分离的细胞能于体外自我更新和分化为成熟的细胞 ,证实为神经前体细胞。

Objective To isolate neural precursor cells from human embryonic cortical tissue of 20 weeks and identify their proliferation capacity as well as differentiating potential. Methods Human fetuses of about 20 weeks from spontaneous abortion were adopted. A serum free medium containing basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) was used to make the neural precursor cell divide continuously in the culture. The proliferating ability of these cells was fested with BrdU. Growth factors were remored so as to induce differentiation of the precursor cells. The cell cycle of the cells in the neurosphere was analysed using flow cytometry. Results Cells from human embryonic cortical tissue could be maintained and propagated in the presence of growth factors. Neurospheres generated continually and the cells in the sphere could incorporate into BrdU. Upon differentiation, the precursor cells gave rise to mature neurons, astrocytes and oligodendrocytes. They retained their multilineage potential over repeated passages.The cell cycle analysis indicated that the cells proliferated actively. Conclusion Cells that able to self renew and differentiate into mature cells in culture cam be isolated from human embryonic tissues. They are werified to be neural precursor cells.