摘要
目的 探讨神经干细胞的原代培养及γ 氨基丁酸 (GABA)能神经元的诱导分化。方法 取孕 16dWistar大鼠的胚鼠全脑 ,进行体外培养 ,并对培养的神经干细胞以及诱导分化的GABA能神经元进行鉴定。结果 培养 2 4h后 ,出现 2～ 4个细胞的细胞球。分化 2d后 ,神经球贴壁后伸出细长突起 ,并可和周边神经球伸出的突起连接。神经球周边可见大量散在贴壁的双极或多极细胞。免疫荧光染色可见神经球均有nestin、NF2 0 0、GFAP阳性细胞。取第 3代神经球行GABA能神经元定向分化。分化 2 4h后 ,实验组细胞球贴壁。 3d后 ,实验组细胞球周边有大量散在分布细胞贴壁生长 ,胞体圆形较大 ,有 1～ 2个细长突起。免疫荧光显示 ,实验组周边散在的贴壁细胞多为GAD6 5阳性细胞。GAD6 5阳性细胞分化率实验组 (85 .97± 2 .78) %、对照组 (18.16± 2 .2 9) % ,P <0 .0 1。结论本实验利用寡核苷酸序列特异性阻断了bHLH基因家族的调控因子之一Hes1,解除了其对bHLH的抑制 ,促进了神经干细胞向神经元的分化。实验还发现 ,阻断Hes1后大大提高了神经干细胞向GA BA神经元分化的比率。
Objective To investigate the primary culture of ne ur al stem cells(NSCs) and induction and differentiation of gamma aminobutyric acid (GABA) neurons.Methods The whole brain tissue of embryonic 1 6 d Wistar rat was taken and underwent primary culture. The cultured NSCs were inducted and differentiated GABA neurons were identified.Results After culturing for 24 h, neurospheres containing 2 to 4 cells appeared. Two days' differentiation later, long synapse grew from the wall of neurosp here, which could connect with the synapses from surrounding neurospheres. A great number of bipolar cells or multipolar cells appeared around the neurosphe re. Immunofluorescence stain showed nestin, NF200,GFAP positive cells in neu rospheres. Third generation of neurosphere underwent GABA neurons orientatin g differentiation. After 24 h's differentitation, neurospheres in the experi mental group archored the wall. Three days later, many dispersal cells grew arch ored the wall surrounding neurospheres in the experimental group. The round cell body was large, with 1 or 2 narrow and long synapses. Immunoflurescence stain s howed the cells around neurospheres in the experimental group mostly were GAD65 positive cells. GAD65 positive cell differentiation rate was (85.9±2.78)% in ex perimental group, while (18.16± 2.29)% in control group, P<0.01. Conclusion Oligonucleotide sequences are applied to specifically block Hes1,which is one of the modulators of bHLH gene family, as a result of re lieving its inhibition to bHLH, and promoting the differentiation from N SCs to neurons. The blocking of Hes1 greatly increasing the differentiation rate from NSCs to GABA neurons is found in the experiment.
出处
《中华神经外科疾病研究杂志》
CAS
2005年第1期44-47,共4页
Chinese Journal of Neurosurgical Disease Research
基金
国家自然科学基金资助项目 (30 370 51 2 )
西京医院临床高新技术面上项目资助项目 (XJGX0 30 33M2 5)
关键词
神经干细胞
分化
GABA神经元
Neural stem cells
Differentiation
Gamma aminobutyric acid (GABA) neu rons
