Can manipulation of differentiation conditions eliminate proliferative cells from a population of ES cell-derived forebrain cells?

Neurogenesis (Austin, Tex.) Pub Date : 2016-01-11 eCollection Date: 2016-01-01 DOI:10.1080/23262133.2015.1127311
Sophie V Precious, Claire M Kelly, Nicholas D Allen, Anne E Rosser
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引用次数: 2

Abstract

There is preliminary evidence that implantation of primary fetal striatal cells provides functional benefit in patients with Huntington's disease, a neurodegenerative condition resulting in loss of medium-sized spiny neurons (MSN) of the striatum. Scarcity of primary fetal tissue means it is important to identify a renewable source of cells from which to derive donor MSNs. Embryonic stem (ES) cells, which predominantly default to telencephalic-like precursors in chemically defined medium (CDM), offer a potentially inexhaustible supply of cells capable of generating the desired neurons. Using an ES cell line, with the forebrain marker FoxG1 tagged to the LacZ reporter, we assessed effects of known developmental factors on the yield of forebrain-like precursor cells in CDM suspension culture. Addition of FGF2, but not DKK1, increased the proportion of FoxG1-expressing cells at day 8 of neural induction. Oct4 was expressed at day 8, but was undetectable by day 16. Differentiation of day 16 precursors generated GABA-expressing neurons, with few DARPP32 positive MSNs. Transplantation of day 8 precursor cells into quinolinic acid-lesioned striata resulted in generation of teratomas. However, transplantation of day 16 precursors yielded grafts expressing neuronal markers including NeuN, calbindin and parvalbumin, but no DARPP32 6 weeks post-transplantation. Manipulation of fate of ES cells requires optimization of both concentration and timing of addition of factors to culture systems to generate the desired phenotypes. Furthermore, we highlight the value of increasing the precursor phase of ES cell suspension culture when directing differentiation toward forebrain fate, so as to dramatically reduce the risk of teratoma formation.

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操纵分化条件能否消除胚胎干细胞来源的前脑细胞群中的增殖细胞?
有初步证据表明,原代胎儿纹状体细胞的植入对亨廷顿氏病(一种神经退行性疾病,导致纹状体中等大小的棘神经元(MSN)丢失)患者的功能有益。原代胎儿组织的稀缺性意味着确定可再生细胞来源以获得供体msn非常重要。胚胎干细胞(ES)在化学定义介质(CDM)中主要默认为端脑样前体,提供了潜在的取之不尽的细胞供应,能够产生所需的神经元。我们使用一种ES细胞系,将前脑标记物FoxG1标记在LacZ报告基因上,评估了已知发育因素对CDM悬浮培养中前脑样前体细胞产量的影响。在神经诱导的第8天,添加FGF2而不添加DKK1增加了foxg1表达细胞的比例。Oct4在第8天表达,但在第16天无法检测到。第16天的前体分化产生表达gaba的神经元,很少有DARPP32阳性的msn。将第8天的前体细胞移植到喹啉酸损伤的纹状体中导致畸胎瘤的产生。然而,移植第16天的前体产生的移植物表达神经元标志物包括NeuN, calbindin和parvalbumin,但在移植后6周没有DARPP32。操纵胚胎干细胞的命运需要在培养系统中优化添加因子的浓度和时间,以产生所需的表型。此外,我们强调了增加胚胎干细胞悬浮培养的前体细胞在引导分化走向前脑命运时的价值,从而显著降低畸胎瘤形成的风险。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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