Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat-Gan, Israel.
Exp Neurol. 2010 May;223(1):119-27. doi: 10.1016/j.expneurol.2009.09.019. Epub 2009 Oct 3.
The enhancement of regeneration of damaged axons in both the peripheral and central nervous systems is a widely pursued goal in clinical medicine. Although some of the molecular mechanisms involved in the intrinsic neurite regeneration program have been elucidated, much additional study is required for development of new therapeutics. The majority of studies in the field of axonal regeneration have utilized animal models due to obvious limitations of the accessibility of human neural tissues. Here we describe the use of human embryonic stem cell (hESC)-derived neurons as a novel model for studying neuronal responses to axonal injury. Neurons were generated using PA6 induction and neurites injured in vitro using trituration or laser microdissection. Lesioned neurons re-extended neurites with distinct growth cones. Expression of proteins associated with regeneration were observed in this human in vitro system, including appearance of importin beta1 in processes after neuritomy. Laser-transected hESC-derived neuronal cultures were analyzed for their transcriptional response to injury using Affymetrix expression microarrays. Profound changes in gene expression were observed over a time course of 2 to 24 hours after lesion. The expression of several genes reported to be involved in axonal injury responses in animal models changed following injury of hESC-derived neurons. Thus, hESC-derived neurons may be a useful in vitro model system for mechanistic studies on human axonal injury and regeneration.
增强外周和中枢神经系统受损轴突的再生能力是临床医学中广泛追求的目标。尽管已经阐明了内在轴突再生程序中涉及的一些分子机制，但为了开发新的治疗方法，还需要进行更多的研究。由于人类神经组织的可及性明显受限，该领域的大多数研究都使用了动物模型。在这里，我们描述了使用人胚胎干细胞（hESC）衍生神经元作为研究神经元对轴突损伤反应的新型模型。使用 PA6 诱导产生神经元，并通过研磨或激光微切割在体外损伤轴突。受损的神经元以明显的生长锥重新延伸轴突。在这个人类体外系统中观察到与再生相关的蛋白质的表达，包括在神经切断后过程中出现的导入蛋白β1。使用 Affymetrix 表达微阵列分析激光切割的 hESC 衍生神经元培养物在损伤后的转录反应。在损伤后 2 至 24 小时的时间过程中观察到基因表达的深刻变化。在动物模型中报道与轴突损伤反应有关的几个基因的表达在 hESC 衍生神经元损伤后发生变化。因此，hESC 衍生神经元可能是研究人类轴突损伤和再生的机制的有用体外模型系统。
