胶质源性神经营养因子对神经前体细胞的促存活和分化作用

Survival-promoting and differenting effects of glial-derived neurotrophic factors on neural precursor cells

目的:观察胶质源性神经营养因子在神经前体细胞的存活和分化过程中的作用及其协同因子。方法:实验于2004-02/2005-08在清华大学玉泉医院神经外科完成。健康孕8～11dSD大鼠20只,体质量300～350g。分别取其胎鼠的皮质和中脑细胞,传代后将细胞按照2×105/孔的密度种植,随机分成8组,每组6孔,重复6次。细胞体外培养9d后进行非多巴胺和多巴胺神经元的免疫组织化学染色/荧光检测,观察胶质源性神经营养因子、转化生长因子β1和/或联丁酰基环腺苷单磷酸后对经碱性成纤维生长因子增殖获得的神经前体细胞--皮质前体细胞和中脑前体细胞分化成多巴胺神经元和非多巴胺神经元的作用。结果:①胶质源性神经营养因子能明显增加发育后期的神经前体细胞的分化和存活。②中脑前体细胞比皮质前体细胞更有可能分化成多巴胺神经元。③应用胶质源性神经营养因子及转化生长因子和/或联丁酰基环腺苷单磷酸后处理神经前体细胞,发现它们能促进皮质前体细胞和中脑前体细胞分化的非多巴胺神经元增加2倍。其中中脑前体细胞分化成的多巴胺神经元数增加8倍,皮质前体细胞分化成多巴胺神经元数也有增加,但仅与分化成的非多巴胺神经元增加相平行。结论:合适的营养因子的加入有助于中脑前体细胞或皮质前体细胞中神经球的存活和分裂增殖及分化,胶质源性神经营养因子、转化生长因子β1和联丁酰基环腺苷单磷酸的协同作用能促进多巴胺神经元的晚期发育。

AIM： To observe the effects of glial-derived neurotrophic factors on survival and differentiation of neural precursor cells and its cooperative factors. METHODS： The study was carried out in the Department of Neurosuregy of Yuquan Hospital of Tsinghua University from February 2004 to August 2005. Mesencephalic and cortex cells of 20 healthy pregnant SD rats of 8-1.1 days with the body mass of 300-350 g were obtained and raised at the density of 2×10^5/pore after passage, and then randomly divided into eight groups with 6 pores in each group, repeated for 6 times. Cells were detected after 9-day-culture in vitro with dopaminergic and non-dopaminergic immunofluorescent assay and immunohistochemical stainning. Effects of glial-defived neurotrophic factors, transforming growth factor β1 and/or dibutyryl cyclic adenosine monophosphate （dbcAMP）on neural precursor cells obtained from the proliferation of basic fibroblast growth factor （bFGF）-dopaminergic and non-dopaminergic neurons differentiated from mesencephalic and cortex precursor cells were observed. RESULTS： ①Glial-derived neurotrophic factors can significantly enhance the differentiation and survival of neural precursor cells in metacyclic stage. ②Mesencephalic precursor cells were more possible to differentiated into dopaminergic neurons than cortex precursor cells. ③It was found in the disposal of neural precursor cells that applications of glial-derived neurotrophic factors, transforming growth factor and dbcAMP can promote the differentiation of non-dopaminergic neurons from mesencephalic and cortex precursor cells increased by two times. Of which the number of dopaminergic neurons differentiated from mesencephalic precursor cells increased by 8 times and that of cortexes also increased, which was only equal to the increasing number of differentiated non-dopaminergic neurons. CONCLUSION： Adding of proper nutritional factors is beneficial for survival, division growth and differentiation of neural bulbs in mesencephalic and cortex precursor cells. Synergistic action of glial-derived neurotrophic factors, transforming growth factor and dbcAMP can promote the development of dopaminergic neurons in advanced stage.