Rapid and Efficient Generation of Functional Motor Neurons From Human Pluripotent Stem Cells Using Gene Delivered Transcription Factor Codes
Rapid and Efficient Generation of Functional Motor Neurons From Human Pluripotent Stem Cells Using Gene Delivered Transcription Factor Codes
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DOI：
10.1038/mt.2011.135
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发表时间：
2011-10-01
影响因子：
12.4
通讯作者：
Kaspar, Brian K.
中科院分区：
文献类型：
--
作者：
Hester, Mark E.;Murtha, Matthew J.;Kaspar, Brian K.
干细胞衍生的运动神经元（MN）越来越多地用于体外疾病建模和用于开发脊髓损伤和疾病（诸如脊髓性肌萎缩症（SMA）和肌萎缩性侧索硬化症（ALS））的细胞替代策略。人胚胎干细胞（hESC）分化为MN，这涉及视黄酸（RA）和激活的音刺猬（SHH）途径是低效的，需要长达60天的开发MN的电生理特性。这种延长的分化过程阻碍了hESC的使用，特别是用于高通量筛选。我们评估了RA/SHH分化的hESCs的MN基因表达谱，以确定参与MN发展的限速因素。基于这种分析，我们开发了一种腺病毒基因递送系统，其编码MN诱导转录因子：神经生成素2（Ngn 2）、胰岛-1（Isl-1）和LIM/同源框蛋白3（Lhx 3）。引人注目的是，这些因子的递送在基因递送后11天诱导具有成熟电生理学特性的功能性MN，其中来自hESC和人诱导多能干细胞（hiPSC）的效率>60-70%。与常规分化技术相比，这种定向编程方法显著减少了产生电生理活性MN所需的时间约30天。我们的研究结果进一步证实了使用转录编码从hESC和hiPSC快速有效地产生确定的细胞类型的潜力。2011年3月19日接收; 2011年6月8日接受; 2011年7月19日在线发表。doi：10.1038/mt.2011.135
Stem cell-derived motor neurons (MNs) are increasingly utilized for modeling disease in vitro and for developing cellular replacement strategies for spinal cord injury and diseases such as spinal muscular atrophy (SMA) and amyotrophic lateral sclerosis (ALS). Human embryonic stem cell (hESC) differentiation into MNs, which involves retinoic acid (RA) and activation of the sonic hedgehog (SHH) pathway is inefficient and requires up to 60 days to develop MNs with electrophysiological properties. This prolonged differentiation process has hampered the use of hESCs, in particular for high-throughput screening. We evaluated the MN gene expression profile of RA/SHH-differentiated hESCs to identify rate-limiting factors involved in MN development. Based on this analysis, we developed an adenoviral gene delivery system encoding for MN inducing transcription factors: neurogenin 2 (Ngn2), islet-1 (Isl-1), and LIM/homeobox protein 3 (Lhx3). Strikingly, delivery of these factors induced functional MNs with mature electrophysiological properties, 11-days after gene delivery, with >60-70% efficiency from hESCs and human induced pluripotent stem cells (hiPSCs). This directed programming approach significantly reduces the time required to generate electrophysiologically-active MNs by approximately 30 days in comparison to conventional differentiation techniques. Our results further exemplify the potential to use transcriptional coding for rapid and efficient production of defined cell types from hESCs and hiPSCs. Received 19 March 2011; accepted 8 June 2011; published online 19 July 2011. doi:10.1038/mt.2011.135