人工脑脊液中苯巴比妥钠对培养神经干细胞分化影响的研究
发布时间：2018-11-28 13:22
【摘要】： 目的本实验目的在于建立神经干细胞培养方法,观察人工脑脊液(ACSF)中不同浓度的苯巴比妥钠(PBS)对新生大鼠脑源性神经干细胞分化成熟的影响。 方法从SD新生大鼠大脑分离、培养神经干细胞,采用无血清培养。免疫细胞化学方法检测神经干细胞特征性标志神经上皮干细胞蛋白(Nestin)表达,用5-溴尿嘧啶核苷(BrdU)检测细胞增殖活性。用四甲基偶氮唑盐法(MTT)观察不同浓度人工脑脊液对神经干细胞存活率的影响。不同浓度苯巴比妥钠在神经干细胞分化中的作用采用免疫组化来检测,并采用扫描电镜对分化后的细胞形态进行观察。 结果(1)经过体外培养获得的神经干细胞能够分化成为神经元和神经胶质细胞,具有多向分化能力,同时有很强的自我更新能力;神经球能够表达Nestin和BrdU,传代培养的神经干细胞性能稳定,经过长期培养后能够保持神经干细胞特点。(2)MTT检测显示,≥40%的人工脑脊液能够明显降低神经干细胞的存活率。(3)免疫细胞化学结果显示,随着人工脑脊液中的苯巴比妥钠浓度的增高,诱导分化后的细胞中神经元百分比逐渐降低,神经胶质细胞百分比逐渐升高,当ACSF中的苯巴比妥钠浓度为10μg/ml时,可显著降低NSCs分化为MAP 2染色阳性细胞的百分率(P＜0.01),当ACSF中的苯巴比妥钠浓度为25μg/ml时,可显著增高NSCs分化为GFAP染色阳性细胞的百分率(P＜0.01)。(4)扫描电镜观察实验组诱导分化后的神经元不具备正常神经元行使功能的形态结构。 结论建立了体外培养大鼠神经干细胞的方法,培养的神经干细胞表达Nestin,有自我更新、多向分化的特点。人工脑脊液中的苯巴比妥钠能够抑制神经干细胞向神经元方向分化,促进其向星型胶质方向分化,且降低了分化后的正常神经元比率。
[Abstract]:Objective to establish a culture method of neural stem cells and to observe the effects of different concentrations of phenobarbital sodium (PBS) in artificial cerebrospinal fluid (ACSF) on the differentiation and maturation of brain-derived neural stem cells in neonatal rats. Methods Neural stem cells were isolated from the brain of neonatal SD rats and cultured without serum. Immunocytochemical method was used to detect the expression of neural epithelial stem cell protein (Nestin), and 5-brominouracil nucleoside (BrdU) was used to detect the proliferative activity of neural stem cells (NSCs). The effects of different concentrations of artificial cerebrospinal fluid (CSF) on the survival rate of neural stem cells were observed by (MTT) method. The effects of phenobarbital sodium at different concentrations on the differentiation of neural stem cells were detected by immunohistochemistry and the morphology of differentiated cells was observed by scanning electron microscope (SEM). Results (1) the neural stem cells cultured in vitro were able to differentiate into neurons and glial cells. The neural stem cells cultured by Nestin and BrdU, have stable performance and can maintain the characteristics of neural stem cells after long term culture. (2) MTT detection showed that, (3) Immunocytochemistry showed that the concentration of phenobarbital sodium increased with the increase of the concentration of phenobarbital sodium in artificial cerebrospinal fluid. The percentage of neurons in the cells induced by differentiation gradually decreased, and the percentage of glial cells increased gradually. When the concentration of phenobarbital sodium in ACSF was 10 渭 g/ml, The percentage of NSCs differentiated into MAP 2 positive cells was significantly decreased (P < 0. 01). When the concentration of phenobarbital sodium in ACSF was 25 渭 g/ml, The percentage of NSCs differentiated into GFAP staining positive cells was significantly increased (P < 0. 01). (4). SEM observation showed that the differentiated neurons in the experimental group did not have the function of normal neurons. Conclusion the method of cultured rat neural stem cells in vitro was established. The cultured neural stem cells expressed Nestin, with the characteristics of self-renewal and multi-differentiation. Phenobarbital sodium in artificial cerebrospinal fluid can inhibit the neural stem cells to differentiate into neurons, promote the differentiation to astrocytes, and decrease the ratio of normal neurons after differentiation.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R329
本文编号：2362939
[Abstract]:Objective to establish a culture method of neural stem cells and to observe the effects of different concentrations of phenobarbital sodium (PBS) in artificial cerebrospinal fluid (ACSF) on the differentiation and maturation of brain-derived neural stem cells in neonatal rats. Methods Neural stem cells were isolated from the brain of neonatal SD rats and cultured without serum. Immunocytochemical method was used to detect the expression of neural epithelial stem cell protein (Nestin), and 5-brominouracil nucleoside (BrdU) was used to detect the proliferative activity of neural stem cells (NSCs). The effects of different concentrations of artificial cerebrospinal fluid (CSF) on the survival rate of neural stem cells were observed by (MTT) method. The effects of phenobarbital sodium at different concentrations on the differentiation of neural stem cells were detected by immunohistochemistry and the morphology of differentiated cells was observed by scanning electron microscope (SEM). Results (1) the neural stem cells cultured in vitro were able to differentiate into neurons and glial cells. The neural stem cells cultured by Nestin and BrdU, have stable performance and can maintain the characteristics of neural stem cells after long term culture. (2) MTT detection showed that, (3) Immunocytochemistry showed that the concentration of phenobarbital sodium increased with the increase of the concentration of phenobarbital sodium in artificial cerebrospinal fluid. The percentage of neurons in the cells induced by differentiation gradually decreased, and the percentage of glial cells increased gradually. When the concentration of phenobarbital sodium in ACSF was 10 渭 g/ml, The percentage of NSCs differentiated into MAP 2 positive cells was significantly decreased (P < 0. 01). When the concentration of phenobarbital sodium in ACSF was 25 渭 g/ml, The percentage of NSCs differentiated into GFAP staining positive cells was significantly increased (P < 0. 01). (4). SEM observation showed that the differentiated neurons in the experimental group did not have the function of normal neurons. Conclusion the method of cultured rat neural stem cells in vitro was established. The cultured neural stem cells expressed Nestin, with the characteristics of self-renewal and multi-differentiation. Phenobarbital sodium in artificial cerebrospinal fluid can inhibit the neural stem cells to differentiate into neurons, promote the differentiation to astrocytes, and decrease the ratio of normal neurons after differentiation.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2007
【分类号】：R329
【参考文献】
相关期刊论文 前5条
1 黄其林,张可成,蔡文琴;脑穿刺伤灶愈合过程中大胶质细胞的变化[J];基础医学与临床;2001年02期
2 李杏色;柴锡庆;;卡马西平对大鼠海马星形胶质细胞形态和GFAP表达的影响[J];基础医学与临床;2006年10期
3 曾可斌,王学峰,晏勇;脑脊液对体外培养骨髓基质细胞生长的影响[J];现代神经疾病杂志;2003年02期
4 卞林,惠国桢,陆华,蒋云召,苗宗宁,王玲,蒋震伟;正常成人脑脊液诱导人胚来源的神经干细胞分化的实验研究[J];实用临床医药杂志;2003年03期
5 笱玉兰,朱遂强,徐金枝,许峰,张广慧,李卫华,阮旭中;匹罗卡品诱导癫痫大鼠内源性神经干细胞的可塑性改变[J];卒中与神经疾病;2005年05期
，本文编号：2362939
本文链接：https://www.wllwen.com/yixuelunwen/binglixuelunwen/2362939.html
最近更新
教材专著
