| Isolation of Mouse Cardiac Neural Crest Cells and Their Differentiation into Smooth Muscle Cells
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Author:
Date:
2017-09-05
[Abstract] Cardiac neural crest cells (CNCCs) originate at the dorsal edge of the neural tube between the otic pit and the caudal edge of the 3rd somite, and migrate into the pharyngeal arches and the heart. We have shown that fibronectin (Fn1) plays an important role in the development of the CNCC by regulating the differentiation of CNCCs into vascular smooth muscle cells around pharyngeal arch arteries (Wang and Astrof, 2016). This protocol describes the isolation of CNCCs from the neural tube and from the caudal pharyngeal arches, and the differentiation of neural crest-derived cells into smooth muscle cells. This protocol was adapted from (Newgreen and Murphy, 2000; Pfaltzgraff et al., 2012).
[摘要] 心脏神经嵴细胞（CNCC）起源于神经管的背部边缘，位于第3个体节的耳穴和尾缘之间，并迁移到咽弓和心脏。 我们已经表明，纤连蛋白（Fn1）通过调节CNCCs到咽弓动脉周围的血管平滑肌细胞的分化，在CNCC的发展中起重要作用（Wang and Astrof，2016）。 该方案描述了CNCC与神经管和尾尾弓的分离，以及神经嵴衍生细胞分化成平滑肌细胞。 该方案从（Newgreen和Murphy，2000; Pfaltzgraff等人，2012）改编。
【背景】以前发表的方案描述了从神经管分离神经嵴细胞。然而，在耳孔和第三体细胞之间的神经管区域中的神经嵴细胞包括有助于许多不同细胞类型的神经嵴细胞群体;例如，迷走神经嵴细胞也来自该区域。在该方案中，我们修改了用于分离心脏神经嵴细胞的常规方法。而不是使用神经管，我们在胚胎期（E）9.5（22-25个体节期）使用尾部咽部弓形区。这是在将心脏神经嵴细胞分化为血管平滑肌细胞之前。神经嵴培养物通常含有污染性间充质细胞，通常表达平滑肌基因。为了鉴定神经嵴衍生细胞，我们从以下交叉产生的胚胎中分离出神经嵴细胞：Fn1flox / flox; ROSAmTmG / mTmG雌性小鼠×Fn1 +/-;Tfap2αIRESCre/ ...
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| Isolation and Culture of Neurospheres for the Study of Pathogenesis of Prion Disease
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Author:
Date:
2014-03-20
[Abstract] Neurosphere contains neural stem cells that are capable of self-renewal and multilineage differentiation including neurons, astrocytes, and oligodendrocytes (Gage, 2000). Cell culture model using differentiated neurosphere cultures are suggested to be a valuable tool for studying the pathogenesis of prion disease at the cellular level (Iwamaru et al., 2013). This protocol describes the procedure for a culture of whole brain-derived neurospheres from newborn mouse brains. Neurosphere formation steadily occurs within a week from the cultures of neonatal whole brains and these cells have stem cell properties.
[摘要] 神经球包含能够自我更新和多向分化的神经干细胞，包括神经元，星形胶质细胞和少突胶质细胞（Gage，2000）。 使用分化的神经球培养物的细胞培养模型被认为是用于研究朊病毒疾病在细胞水平的发病机制的有价值的工具（Iwamaru等人，2013）。 该协议描述了来自新生小鼠脑的全脑衍生的神经球的培养的程序。 神经球形成稳定地发生在新生儿全脑的培养物的一周内，并且这些细胞具有干细胞特性。
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| Neural Stem Cell Differentiation and Prion Infection
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Author:
Date:
2014-03-20
[Abstract] Prion diseases are transmissible, fatal, neurodegenerative diseases in human and animals. The molecular basis of neurodegeneration in prion diseases is largely unclear. Developing a cellular model capable of monitoring prion-induced cytotoxicity would be a promising approach for better understanding the prion pathogenesis. One candidate cellular assay is a model based on neurospheres, which contains neural stem cells (NSCs). Both undifferentiated and differentiated NSCs have been demonstrated to be permissive to prion infection, and prion-induced cytopathic changes in differentiated neruosphere cultures were reported (Iwamaru et al., 2013). This protocol describes the procedure to induce differentiation of NSCs from transgenic mice overexpressing prion protein (tga20 mice) into ...
[摘要] 朊病毒疾病是人类和动物中可传播的，致命的，神经变性疾病。 朊病毒疾病中神经变性的分子基础很大程度上不清楚。 开发能够监测朊病毒诱导的细胞毒性的细胞模型将是更好地理解朊病毒发病机制的有前途的方法。 一种候选细胞测定是基于神经球的模型，其含有神经干细胞（NSC）。 未分化和分化的NSCs都被证明是容许朊病毒感染，并且报道了分化的神经球培养物中的朊病毒诱导的细胞病变变化（Iwamaru等人，2013）。 该协议描述了诱导NSCs从过度表达朊病毒蛋白（tga20小鼠）的转基因小鼠分化成易感染朊病毒感染的培养物的过程。
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