Characterization of neurons from immortalized dental pulp stem cells for the study of neurogenetic disorders.
Characterization of neurons from immortalized dental pulp stem cells for the study of neurogenetic disorders.
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从永生化的牙髓干细胞中的神经元来研究神经遗传疾病。
DOI：
10.1016/j.scr.2015.11.004
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发表时间：
2015-11
影响因子：
1.2
通讯作者：
Reiter LT
中科院分区：
文献类型：
--
作者：
Urraca N;Memon R;El-Iyachi I;Goorha S;Valdez C;Tran QT;Scroggs R;Miranda-Carboni GA;Donaldson M;Bridges D;Reiter LT
神经遗传综合征研究和治疗的一个主要挑战是获取受影响个体的活神经元进行研究。尽管目前有多种干细胞来源，但获取这些干细胞涉及侵入性操作，生成可能很困难或昂贵，而且数量有限。牙髓干细胞（DPSC）是驻留在脱落牙髓深处的多能干细胞。为了研究 DPSC 的特征，使其成为转化研究的宝贵资源，我们对对照 DPSC 和衍生神经元进行了一系列活力、衰老、永生化和基因表达研究。我们调查了初级 DPSC 的基本运输条件和最大通过次数。我们使用人端粒酶逆转录酶 (hTERT) 永生化对照 DPSC，并通过 RNA-seq 评估神经元分化潜力和全局基因表达变化。我们发现永生化 DPSC 的神经元与非永生化 DPSC 具有相同的形态和电生理特性。我们还表明，DPSC 分化为神经元会显着改变 1305 转录本的基因表达。在这里，我们表明基因表达的这些变化与转录抑制因子 REST/NSRF 的蛋白质水平的变化同时发生，已知 REST/NSRF 参与神经元分化。永生化显着改变了神经元分化后 183 个基因的表达，其中 94 个基因在分化过程中也发生了变化。我们的研究表明，可以从保存≥72小时的牙齿中获得活的DPSC，然后可以将其永生化，并且仍然产生用于体外研究的功能性神经元，但是组成型hTERT永生化并不是长期使用源自患者的DPSC进行疾病研究的最佳方法。
A major challenge to the study and treatment of neurogenetic syndromes is accessing live neurons for study from affected individuals. Although several sources of stem cells are currently available, acquiring these involve invasive procedures, may be difficult or expensive to generate and are limited in number. Dental pulp stem cells (DPSC) are multipotent stem cells that reside deep the pulp of shed teeth. To investigate the characteristics of DPSC that make them a valuable resource for translational research, we performed a set of viability, senescence, immortalization and gene expression studies on control DPSC and derived neurons. We investigated the basic transport conditions and maximum passage number for primary DPSC. We immortalized control DPSC using human telomerase reverse transcriptase (hTERT) and evaluated neuronal differentiation potential and global gene expression changes by RNA-seq. We show that neurons from immortalized DPSC share morphological and electrophysiological properties with non-immortalized DPSC. We also show that differentiation of DPSC into neurons significantly alters gene expression for 1305 transcripts. Here we show that these changes in gene expression are concurrent with changes in protein levels of the transcriptional repressor REST/NSRF, which is known to be involved in neuronal differentiation. Immortalization significantly altered the expression of 183 genes after neuronal differentiation, 94 of which also changed during differentiation. Our studies indicate that viable DPSC can be obtained from teeth stored for ≥72hrs, these can then be immortalized and still produce functional neurons for in vitro studies, but that constitutive hTERT immortalization is not be the best approach for long term use of patient derived DPSC for the study of disease.