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目的建立成年大鼠嗅球神经干细胞分离培养和鉴定方法,探索新的成年神经干细胞种子来源。方法用无血清方法分离培养成年大鼠嗅球来源的神经干细胞;用克隆培养、BrdU整合的方法检验培养细胞的干细胞特性;用免疫荧光细胞化学的方法检测BrdU、神经干细胞标记物Nestin和SOX2,分化的细胞标记物Tuj1、GFAP、NG2的表达。结果从成年大鼠嗅球能够分离、培养出具有自我更新、增殖的神经球,构成神经球的细胞呈Nestin和SOX2阳性,它们分化后产生Tuj1阳性的神经元、GFAP阳性的星形胶质细胞、NG2阳性的少突胶质细胞。结论成年大鼠嗅球存在神经干细胞,能够在体外进行培养、增殖、分化,是神经干细胞的新的种子来源。
OBJECTIVE: To establish a method for the isolation, culture and identification of neural stem cells from olfactory bulb in adult rats and explore the origin of new adult neural stem cells. Methods The neural stem cells derived from the olfactory bulb of adult rats were isolated and cultured by serum-free method. The characteristics of stem cells in cultured cells were detected by Clonculture and BrdU integration. Nestin and SOX2, markers of BrdU and neural stem cells, were detected by immunofluorescence cytochemistry. Of cell markers Tuj1, GFAP, NG2. Results Neurospheres with self-renewal and proliferation were isolated and cultured from the olfactory bulb of adult rats. Nestin and SOX2-positive neurons were found in neurospheres. After differentiation, they produced Tuj1-positive neurons, GFAP-positive astrocytes, NG2-positive oligodendrocytes. Conclusion There are neural stem cells in the olfactory bulb of adult rats, which can be cultured, proliferated and differentiated in vitro. It is a new source of seeds for neural stem cells.