Direct Conversion of Equine Adipose-Derived Stem Cells into Induced Neuronal Cells Is Enhanced in Three-Dimensional Culture
Direct Conversion of Equine Adipose-Derived Stem Cells into Induced Neuronal Cells Is Enhanced in Three-Dimensional Culture
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DOI：
10.1089/cell.2015.0046
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发表时间：
2015-12-01
影响因子：
1.6
通讯作者：
Strappe, Padraig M.
中科院分区：
文献类型：
--
作者：
Petersen, Gayle F.;Hilbert, Bryan J.;Strappe, Padraig M.
从马身上培养神经元的能力可能会让我们对马的神经系统疾病进行进一步的研究。在这项研究中，我们证明了使用神经元转录因子Brn 2、Ascl 1、Myt 11（BAM）和NeuroD 1的慢病毒载体表达和确定的化学诱导培养基的组合从马脂肪源干细胞（EADSC）诱导神经元细胞的产生，III-微管蛋白阳性诱导的神经元细胞显示出圆形和紧凑的细胞体的独特神经元形态，广泛的神经突生长，和过程的分支。此外，我们研究了维度对神经元转分化的影响，比较传统的二维（2D）单层培养与三维（3D）培养的多孔聚苯乙烯支架。在三维培养中，神经元转分化得到增强，细胞均匀分布在支架表面和整个支架上。在3D培养中转分化效率增加，与2D培养相比，平均转化百分比增加超过100%。此外，诱导的神经元细胞被证明通过巢蛋白阳性前体状态，与MAP 2和突触蛋白2的表达显着增加，在3D培养。这些发现将有助于增加我们对马神经发病机制的理解，在疾病建模，药物筛选和细胞替代治疗马神经系统疾病方面具有前瞻性作用。
The ability to culture neurons from horses may allow further investigation into equine neurological disorders. In this study, we demonstrate the generation of induced neuronal cells from equine adipose-derived stem cells (EADSCs) using a combination of lentiviral vector expression of the neuronal transcription factors Brn2, Ascl1, Myt1l (BAM) and NeuroD1 and a defined chemical induction medium, with III-tubulin-positive induced neuronal cells displaying a distinct neuronal morphology of rounded and compact cell bodies, extensive neurite outgrowth, and branching of processes. Furthermore, we investigated the effects of dimensionality on neuronal transdifferentiation, comparing conventional two-dimensional (2D) monolayer culture against three-dimensional (3D) culture on a porous polystyrene scaffold. Neuronal transdifferentiation was enhanced in 3D culture, with evenly distributed cells located on the surface and throughout the scaffold. Transdifferentiation efficiency was increased in 3D culture, with an increase in mean percent conversion of more than 100% compared to 2D culture. Additionally, induced neuronal cells were shown to transit through a Nestin-positive precursor state, with MAP2 and Synapsin 2 expression significantly increased in 3D culture. These findings will help to increase our understanding of equine neuropathogenesis, with prospective roles in disease modeling, drug screening, and cellular replacement for treatment of equine neurological disorders.