Functional surfaces for efficient differentiation of neural stem/progenitor cells into dopaminergic neurons.
Functional surfaces for efficient differentiation of neural stem/progenitor cells into dopaminergic neurons.
复制标题
用于将神经干/祖细胞有效分化为多巴胺能神经元的功能表面。
DOI：
10.1002/jbm.a.36602
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发表时间：
2019
期刊：
影响因子：
--
通讯作者：
H.
中科院分区：
文献类型：
--
作者：
Nakaji-Hirabayashi;T.: Fujimoto;K.; Yoshikawa;C.; Kitano;H.
神经干细胞/祖细胞向多巴胺能神经元的有效分化已有多种技术和系统的报道。虽然使用这些技术可以获得相对较高比例的多巴胺能神经元，但分化的细胞表现出不同的细胞表型，如星形胶质细胞和少突胶质细胞。产生高纯度的多巴胺能神经元对于基于细胞的治疗和多巴胺能神经元功能的体外评估是重要的。在这项研究中，我们开发了一种用几种神经营养因子和神经元细胞粘附蛋白固定的培养表面，用于神经干/祖细胞向多巴胺能神经元的有效分化。利用基因工程合成的低聚组氨酸融合脑源性神经营养因子和胶质细胞系源性神经营养因子，通过金属螯合共同固定在表面。为了促进细胞粘附，还将层粘连蛋白衍生的细胞粘附嵌合蛋白（LN‐G）固定在表面。用这些蛋白固定底物培养14天后，大约40%的细胞表达酪氨酸羟化酶（多巴胺能神经元的标记物），分化效率比之前报道的提高了三倍。此外，30天后酪氨酸羟化酶阳性细胞的数量增加到培养物的80%左右。这些细胞分泌多巴胺并表达多巴胺能神经元特异性基因。有趣的是，除了神经细胞（未成熟和成熟的多巴胺能神经元）外，蛋白质锚定表面未检测到细胞类型（胶质细胞和少突胶质细胞）。我们的研究结果表明，高纯度的多巴胺能神经元可以使用新的底物获得，而无需额外的纯化步骤，如细胞分选。©2018 Wiley期刊公司中国生物医学工程学报，2016,31(1):444 - 444。
Various techniques and systems have been reported for the efficient differentiation of neural stem/progenitor cells into dopaminergic neurons. Although a comparatively high percentage of dopaminergic neurons can be obtained using these techniques, the differentiated cells display varied cellular phenotypes such as astrocytes and oligodendrocytes. Generation of highly pure dopaminergic neurons is important for cell‐based therapy and in vitro evaluation of dopaminergic neuron function. In this study, we developed a culture surface anchored with several neurotrophic factors and a neuronal cell‐adhesive protein for efficient differentiation of neural stem/progenitor cells into dopaminergic neurons. Oligohistidine‐fused brain‐derived neurotrophic factor and glial cell line–derived neurotrophic factor, synthesized using genetic engineering, were co‐immobilized on the surface via metal chelation. To facilitate cell adhesion, a cell‐adhesive chimeric protein derived from laminin (LN‐G) was also immobilized on the surface. Approximately 40% of the cells cultured for 14 days with these protein‐immobilized substrates expressed tyrosine hydroxylase, a marker of dopaminergic neurons, with a three‐fold increase in differentiation efficiency than that reported previously. In addition, the number of tyrosine hydroxylase‐positive cells increased to approximately 80% of the culture after 30 days. These cells secreted dopamine and expressed dopaminergic neuron‐specific genes. Interestingly, cell types (glial cells and oligodendrocytes) other than neuronal cells (immature and mature dopaminergic neurons) were not detected on the protein‐anchored surface. Our results demonstrate that highly pure dopaminergic neurons can be exclusively obtained using the novel substrate without extra purification steps such as cell sorting. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 860–871, 2019.