Axon Regeneration in Vitro on Physiologically Relevant Substrata

D. Shewan, K. Bedi, M. Berry, J. Winter, James Cohen
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引用次数: 8

Abstract

The study of axon growth in culture is limited by a poor understanding of the relative contribution of each of a complex array of factors, which include diffusible, axon growth-modulating molecules and substrate-bound guidance cues available to developing and regenerating neurons in vivo. With the objective of more closely mimicking in vivo conditions, one approach we have exploited employs thin cryosections of appropriate regions of unfixed nervous tissue as culture substrata for the growth of regenerating neurons. By using this technique it is possible to culture different populations of neurons on substrata in which environmental growth-modulating factors are preserved. This form of bioassay has facilitated the study of the different neurite outgrowth responses of neurons both from different sources and at different developmental ages on varying native substrata. Using this method we have demonstrated that mature dorsal root ganglion neurons (DRG) will regrow axons only on predegenerated sciatic nerve in vitro, while immature DRG extend neurites on both intact and degenerated sciatic nerve. In contrast, both mature and neonatal DRG fail to regenerate on either fully myelinated mature optic nerve or unmyelinated embryonic optic nerve. Moreover, neonatal retinal ganglion cells do not regenerate on any of these substrata.
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轴突在生理相关基质上的体外再生
对培养轴突生长的研究受到对一系列复杂因素的相对贡献理解不足的限制,这些因素包括可扩散的轴突生长调节分子和可用于体内神经元发育和再生的底物结合引导线索。为了更接近地模拟体内条件,我们采用了一种方法,将不固定的神经组织的适当区域薄切片作为再生神经元生长的培养基质。通过使用这种技术,可以在保存环境生长调节因子的基质上培养不同的神经元群体。这种形式的生物测定有助于研究不同来源和不同发育年龄的神经元在不同天然基质上的不同神经突生长反应。利用这种方法,我们证明了成熟的背根神经节神经元(DRG)只能在体外预变性的坐骨神经上再生轴突,而未成熟的DRG在完整和变性的坐骨神经上都能延伸神经突。相比之下,成熟和新生儿DRG在完全有髓鞘的成熟视神经或无髓鞘的胚胎视神经上都不能再生。此外,新生儿视网膜神经节细胞不能在任何这些基质上再生。
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