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目的 探讨人脐血单个核细胞 (MNCs)体外培养后的神经元特有分子表达和形态特征。 方法 密度梯度离心法分离脐血中单个核细胞 ,部分接种于培养瓶内 ,部分细胞接种在 6孔培养板内。倒置显微镜下观察培养细胞形态变化。RT PCR方法检测MNCs培养前后神经细胞标志物nestin、NF M及MAP2mRNA的表达 ,培养 14d后行免疫细胞化学鉴定。 结果 培养前脐血MNCs胞体小呈圆形 ,神经干 前体细胞特异性抗体nestin阳性细胞、神经元特异性抗体MAP2和NF M阳性细胞散在分布 ,阳性率分别为 1 5 %、3 4 %和 2 5 %。培养 14d后 ,倒置显微镜下可见一些有多个粗长突起的细胞群 ,相邻细胞突起连成网状 ;免疫细胞化学检测MAP2、NF M染色阳性细胞成片状分布 ,阳性率分别为 2 7 6 %和 2 1 7%。RT PCR检测脐血单个核细胞nestin、NF M及MAP2基因表达呈阳性 ,培养 14d后上述基因表达上调。 结论 脐血细胞中可能沉寂有神经 (干 )前体细胞 ,经体外培养后能分化为具有一定形态的类神经细胞。
Objective To investigate the neuron-specific molecular expression and morphological characteristics of human cord blood mononuclear cells (MNCs) cultured in vitro. Methods Mononuclear cells in cord blood were isolated by density gradient centrifugation, partially in culture flasks and partially seeded in 6-well plates. The morphological changes of cultured cells were observed under inverted microscope. RT-PCR was used to detect the expression of nestin, NF M and MAP2mRNA in neural cell line before and after culture of MNCs, and the expression of nestin, NF M and MAP2mRNA was detected by immunocytochemistry 14 days after culture. Results The cell bodies of cord blood MNCs were small and round before transplantation. The numbers of nestin positive cells, neuron specific antibodies MAP2 and NF M positive cells were 15% and 34% respectively 2 5%. After culturing for 14 days, some cell groups with many protuberant protuberances were observed under an inverted microscope. The adjacent cell protuberances reticulated. Immunocytochemistry detected the positive distribution of MAP2 and NF M positive cells in the form of flake. The positive rates were 27 6% and 21.7%. RT-PCR detection of cord blood mononuclear cells nestin, NF M and MAP2 gene expression was positive after 14 days of culture the above genes were up-regulated. Conclusion Umbilical cord blood cells may have neural precursors (stem) precursors quietly, which can differentiate into certain types of neural cells after in vitro culture.