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背景:神经干细胞已被应用于治疗成人颅脑损伤及其他神经系统退行性疾病中。体外培养的神经干细胞脱离了体内正常的生理环境,所产生的反应是否与在生理条件下一致?诱导内源性的神经干细胞增殖分化可能是为实践开辟出另一条更有前程的途径。目的:探讨不同胎龄胎脑皮质额叶神经干细胞的发育规律。设计:随机实验观察。单位:南方医科大学附属珠江医院儿科。对象:收集2003-10/2004-03南方医科大学附属珠江医院妇产科胎龄16～36周的自愿水囊引产胎儿90例,取其皮质额叶脑组织作为实验材料。胎儿母亲均查体健康,水囊引产胎儿经家属及胎儿母亲签字同意、并经南方医科大学主管部门同意后进行实验。按胎龄16,20,24,28,32,36周分为6组,每组15例。方法:水囊引产胎儿娩出后在无菌条件下进行处理,打开头皮及颅骨,撕除脑表面被膜,切取皮质额叶脑组织,进行固定后切片,采用免疫组织化学及光镜技术对人胎脑皮质额叶神经干细胞的分布、形态、种类、生长方式以及数量进行检测。光学显微镜下观察细胞的形态及阳性表达情况。用替代法作阴性对照实验。在光学显微镜×400下观察,Nestin阳性细胞胞浆染成棕黄色,判定为神经干细胞,每组每个样本取2张载玻片,每张载玻片选10个视野,每组计算镜下300个视野的总细胞数和阳性细胞数。计算每组神经干细胞阳性检出率。主要观察指标:不同胎龄组人胎脑皮质额叶Nestin蛋白阳性神经干细胞的形态学及数量发育特征。结果:人胎脑皮质额叶存在神经干细胞,主要分布在锥体细胞层及内颗粒细胞层,呈小圆形及椭圆形,以前者最多,胞核相对较大,空泡状,1～3个核仁不等,大部分细胞染色质疏松,胞浆表达明显,有的圆形细胞呈僧帽状,僧帽细胞可见短突起,椭圆形细胞有2个突起,神经干细胞呈明显的区域性分布,其中有由数个细胞形成的神经干细胞集落,似体外培养形成的神经干细胞克隆球,偶见对称性分裂的神经干细胞。不同胎龄组16,20,24,28,32,36周组人胎脑皮质额叶Nestin蛋白阳性神经干细胞检出率逐渐减少(15.59%,13.48%,11.62%,10.52%,9.87%,6.68%,χ2=1265.152,P<0.01)。结论:不同胎龄人胎脑皮质额叶神经干细胞在分布、形态、种类、生长方式及数量上存在一定的差异和自身的发育特征,其数量随着胎龄的增加而减少。
Background: Neural stem cells have been used to treat craniocerebral injury in adults and other neurodegenerative diseases. In vitro cultured neural stem cells from the body’s normal physiological environment, the response is consistent with the physiological conditions? Induction of proliferation and differentiation of endogenous neural stem cells may be opened to practice for another more promising way. Objective: To investigate the developmental rules of neural stem cells in frontal cortex of different gestational age fetuses. Design: Randomized experimental observation. Unit: Zhujiang Hospital affiliated to Southern Medical University. PARTICIPANTS: Totally 90 fetuses with gestational age of 16 to 36 weeks in Department of Obstetrics and Gynecology of Zhujiang Hospital Affiliated to Southern Medical University were collected from 2003-10 / 2004-03, and their cortical frontal lobe brain tissue was taken as experimental material. Fetus and mothers were physical examination, fetus induced by the fetus and the fetus mother signed the consent of the South Medical University department agreed with the consent of the experiment. By gestational age 16,20,24,28,32,36 weeks were divided into 6 groups, 15 cases in each group. Methods: After delivery of fetus, the fetus was treated under aseptic condition, the scalp and skull were opened, the surface of the brain was removed and the cortex of the frontal lobe brain was dissected. After fixation, the fetus was fixed and stained with immunohistochemistry and light microscope Cortical frontal nerve stem cells distribution, morphology, type, growth pattern and the number of tests. The morphology and the positive expression of the cells were observed under an optical microscope. Alternative method for negative control experiments. Observed in optical microscope × 400, Nestin-positive cells were stained brown, cytoplasm stained for neural stem cells, each sample taken two slides, each slide selected 10 fields, each calculated under the microscope Total cell number and number of positive cells in 300 fields. Calculate the positive detection rate of each group of neural stem cells. MAIN OUTCOME MEASURES: Morphology and quantitative characteristics of Nestin positive neural stem cells in frontal cortex of human fetal cortex in different gestational age groups. Results: There were neural stem cells in the frontal cortex of human fetal brain, which mainly distributed in the pyramidal cell layer and the inner granular cell layer. They were small round and oval, the former had the most, the nucleus was relatively large, vacuolar, 1 ~ 3 A nucleolus range, most of the cells loose chromatin, cytoplasm obvious, some round cells were monk-shaped, monkshood cells can be seen short prominence, oval cells have 2 protrusions, neural stem cells showed a clear regional Distribution, of which there are several cells formed by the neural stem cell colonies, like in vitro culture of the formation of neural stem cell clone ball, occasionally symmetrical division of neural stem cells. The detection rate of Nestin protein positive neural stem cells in the frontal cortex of human fetus in different gestational age groups decreased gradually at 16, 20, 24, 28, 32, 36 weeks (15.59%, 13.48%, 11.62%, 10.52%, 9.87%, 6.68 %, χ2 = 1265.152, P <0.01). CONCLUSION: There are some differences in the distribution, morphology, type, growth pattern and quantity of neural stem cells in frontal cortex of different gestational age human fetuses and their developmental features. The number of neural stem cells decreases with the increase of gestational age.