Differentiation of the SH-SY5Y Human Neuroblastoma Cell Line.
Differentiation of the SH-SY5Y Human Neuroblastoma Cell Line.
复制标题
SH-SY5Y人神经母细胞瘤细胞系的分化。
DOI：
10.3791/53193
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发表时间：
2016-02-17
期刊：
影响因子：
--
通讯作者：
Szpara ML
中科院分区：
文献类型：
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作者：
Shipley MM;Mangold CA;Szpara ML
拥有合适的体内和体外系统，为人类疾病提供翻译模型，是神经生物学和神经科学研究的一个不可或缺的方面。用于神经生物学的传统体外实验模型包括大鼠和小鼠的原代神经元培养，神经母细胞瘤细胞系包括大鼠B35和小鼠Neuro-2A细胞，大鼠PC12细胞，以及短期切片培养。虽然许多研究人员依赖于这些模型，但它们缺乏人类的组成部分，所观察到的实验效应可能是各自物种独有的，可能不会在人类身上发生相同的情况。此外，尽管这些细胞是神经元，但它们的核型可能不稳定，这使得它们的使用在基因表达研究和细胞信号转导的可重复性研究中存在问题。因此，开发更一致的人类神经系统疾病模型是很重要的。通过对染色体稳定的人神经母细胞瘤细胞系SH-SY5Y进行分化，以下过程描述了一种易于遵循、可重复性的方法，以获得同质和可存活的人类神经元培养。这种方法集成了前面描述的几种方法，并基于从介质中顺序去除血清。时间线包括逐渐的血清饥饿，引入细胞外基质蛋白和神经营养因子。这允许神经元分化，同时选择上皮细胞作为对照，从而产生同质的神经元培养。具有代表性的结果表明，SH-SY5Y神经母细胞瘤细胞成功地从最初的上皮样细胞表型分化为更具扩张性和分支的神经元表型。该协议提供了一种可靠的方法来生成可用于后续生化和分子分析的神经元培养的同质群体，这为研究人员提供了更准确的人类感染和疾病的翻译模型。
Having appropriate in vivo and in vitro systems that provide translational models for human disease is an integral aspect of research in neurobiology and the neurosciences. Traditional in vitro experimental models used in neurobiology include primary neuronal cultures from rats and mice, neuroblastoma cell lines including rat B35 and mouse Neuro-2A cells, rat PC12 cells, and short-term slice cultures. While many researchers rely on these models, they lack a human component and observed experimental effects could be exclusive to the respective species and may not occur identically in humans. Additionally, although these cells are neurons, they may have unstable karyotypes, making their use problematic for studies of gene expression and reproducible studies of cell signaling. It is therefore important to develop more consistent models of human neurological disease. The following procedure describes an easy-to-follow, reproducible method to obtain homogenous and viable human neuronal cultures, by differentiating the chromosomally stable human neuroblastoma cell line, SH-SY5Y. This method integrates several previously described methods and is based on sequential removal of serum from media. The timeline includes gradual serum-starvation, with introduction of extracellular matrix proteins and neurotrophic factors. This allows neurons to differentiate, while epithelial cells are selected against, resulting in a homogeneous neuronal culture. Representative results demonstrate the successful differentiation of SH-SY5Y neuroblastoma cells from an initial epithelial-like cell phenotype into a more expansive and branched neuronal phenotype. This protocol offers a reliable way to generate homogeneous populations of neuronal cultures that can be used for subsequent biochemical and molecular analyses, which provides researchers with a more accurate translational model of human infection and disease.