摘要
目的观察Notch-1信号在骨髓间质干细胞(MSC)分化为神经细胞中的作用。方法采用RNA干扰技术,构建小鼠Notch-1shRNA,转染小鼠MSC后诱导其分化为神经细胞;同时以未转染MSC和转染甘油醛-3-磷酸脱氢酶(GAPDH)shRNA为对照。采用免疫细胞化学染色和Western印迹检测巢蛋白、神经元特异性烯醇化酶(NSE)、神经微丝蛋白200(NF200)、胶质纤维酸性蛋白(GFAP)和Notch-1蛋白表达;原位末端标记染色评价细胞凋亡率。结果诱导6d后,转染Notch-1shRNA诱导后细胞形成较典型的神经细胞形态,同时高效表达巢蛋白、NSE和NF200等,不表达GFAP,对照组无此现象。但是转染mNotch-1shRNA后细胞凋亡率(13·3%±2·3%)显著高于对照组。结论MSC的横向分化过程可能与神经干细胞类似,阻断Notch信号通路,会增加MSC向神经细胞分化的效果。
Objective To investigate the role of Notch-1 signaling in bone marrow mesenchymal stem ceils （MSCs） differentiating into neurons. Methods Mice Notch-1 small hairpin RNA （ mNotch-1 shRNA） was constructed and transfected into the MSCs obtained from the tibiae of BALB/c mice. MSCs transfected with glyseraldehyde-3-phosphate hydroxygenase （GADPH） shRNA and untransfected MSCs were used as controls. The cell survival rate was detected by ELISA. The MSCs of different groups were cultured in Neurobasal-A medium so as to be induced to differentiate into neurons. Apoptosis of the MSCs was detected by TUNEL. Results After induction of 6 days the MSCs transfected with mNotch-1 shRNA displayed typical neuronal morphology and high expression of neuron-specific markers： nestin, neuron-specific enolase （NSE）, neurofilament 200 （NF 200）, and Notch-1 protein, however, gilal fibrillary acidic protein （GFAP）, the glia-specific marker, was not detected. The percentage of apoptotic cells in the MSCs transfected with mNotch-1 shRNA was 13.3% ± 2.3%, significantly higher than those of the MSCs transfected with mGAPH shRNA and untransfected MSCs （4. 7% ± 0. 5% and 4. 5% ± 0. 4%, both P 〈 0. 01 ）. Condusion Block of the Notch signal pathway may increase the differentiation of MSCs into neurons.
出处
《中华医学杂志》
CAS
CSCD
北大核心
2006年第41期2916-2921,共6页
National Medical Journal of China
基金
国家自然科学基金资助项目(30200128
30540062)
中国博士后科学基金资助项目(第29
33期)
关键词
骨髓间质干细胞
神经元
RNA干扰
Marrow mesenchymal stem cells
Neurons
RNA interference
