A low-cost microwell device for high-resolution imaging of neurite outgrowth in 3D.
A low-cost microwell device for high-resolution imaging of neurite outgrowth in 3D.
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DOI：
10.1088/1741-2552/aaaa32
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发表时间：
2018-06
影响因子：
4
通讯作者：
Suter DM
中科院分区：
文献类型：
--
作者：
Ren Y;Mlodzianoski MJ;Lee AC;Huang F;Suter DM
目前的神经元细胞培养大多是在二维表面进行的，缺乏神经元自然环境的许多重要特征，包括地形线索、可变形的细胞外基质以及三维空间各向同性或各向异性。尽管三维(3D)细胞培养系统提供了比2D系统更具生理学相关性的环境，但由于缺乏易于制造和使用的设备，它们的普及受到了极大的阻碍。我们的目标是开发一种广泛适用的3D培养程序，以促进神经元培养从2D过渡到3D。我们制作了一种用于3D神经细胞培养的简单微孔设备，该设备价格低廉，易于组装，并与常用的成像技术完全兼容，包括超分辨率显微镜。我们开发了一种新的凝胶混合物来支持海兔袋细胞神经元的3D轴突再生，该系统已被广泛用于二维生长锥动力学的定量分析。我们发现，在3D培养中，袋状细胞生长锥的形态和生长模式与体内观察到的生长锥非常相似。我们展示了我们的设备能够对细胞骨架和信号蛋白以及细胞器进行高分辨率成像。神经细胞培养一直是神经科学家研究神经元在受控环境中行为的宝贵工具。与2D相比，3D培养的神经元保留了大部分天然特征，同时提供了更高的可达性、可控性和重复性。我们希望我们的微孔装置将有助于更广泛地采用3D神经元培养来研究轴突再生的机制。
Current neuronal cell culture is mostly performed on two-dimensional (2D) surfaces, which lack many of the important features of the native environment of neurons, including topographical cues, deformable extracellular matrix, and spatial isotropy or anisotropy in three dimensions. Although three-dimensional (3D) cell culture systems provide a more physiologically relevant environment than 2D systems, their popularity is greatly hampered by the lack of easy-to-make and -use devices. We aim to develop a widely applicable 3D culture procedure to facilitate the transition of neuronal cultures from 2D to 3D. We made a simple microwell device for 3D neuronal cell culture that is inexpensive, easy to assemble, and fully compatible with commonly used imaging techniques, including super-resolution microscopy. We developed a novel gel mixture to support 3D neurite regeneration of Aplysia bag cell neurons, a system that has been extensively used for quantitative analysis of growth cone dynamics in 2D. We found that the morphology and growth pattern of bag cell growth cones in 3D culture closely resemble the ones of growth cones observed in vivo. We demonstrated the capability of our device for high-resolution imaging of cytoskeletal and signaling proteins as well as organelles. Neuronal cell culture has been a valuable tool for neuroscientists to study the behavior of neurons in a controlled environment. Compared to 2D, neurons cultured in 3D retain the majority of their native characteristics, while offering higher accessibility, control, and repeatability. We expect that our microwell device will facilitate a wider adoption of 3D neuronal cultures to study the mechanisms of neurite regeneration.
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影响因子：
5.7
作者：
Bentley, David;O'Connor, Timothy P.
通讯作者：
O'Connor, Timothy P.
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作者：
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通讯作者：
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影响因子：
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作者：
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通讯作者：
Suter DM