Revisiting nestin expression in retinal progenitor cells in vitro and after transplantation in vivo
Revisiting nestin expression in retinal progenitor cells in vitro and after transplantation in vivo
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DOI：
10.1016/j.exer.2007.01.014
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发表时间：
2007-06-01
影响因子：
3.4
通讯作者：
Sadda, SriniVas R.
中科院分区：
文献类型：
--
作者：
Qiu, Guanting;Seiler, Magdalene J.;Sadda, SriniVas R.
本研究的目的是研究神经外胚层干细胞和反应性神经胶质标记物Nestin与各种成熟的视网膜细胞标记物在体外扩增的视网膜前体细胞(RPC)中的共表达情况，然后进行体外诱导或视网膜下移植。取胚胎第17天的大鼠视网膜，在无血清培养条件下扩增RPC，并用全反式维甲酸(RA)诱导分化。诱导后，对细胞进行巢蛋白染色，结合视网膜神经元和神经胶质标记物。诱导前后收集培养细胞进行定量RT-PCR基因表达分析。在第二个系列中，将第2代RPC移植到出生后第28天的S334ter-3视网膜变性大鼠的视网膜下间隙中。移植后1~4周取材，行巢蛋白和各种视网膜特异性神经元标记物免疫组织化学双重染色。经RA处理的培养的RPC与多种视网膜特异性标记物共表达巢蛋白，包括蛋白激酶Cα(PKC)、神经细丝200(NTF200)、细胞视黄醛结合蛋白(CRALBP)和视紫红质。在RA诱导后，定量RT-PCR分析显示Nestin、PAX-6、THY 1.1表达下调。和PKCα，视紫红质、胶质纤维酸性蛋白(GFAP)和CRX上调。在体内RPC移植中，除了与宿主视网膜中的GFAP阳性细胞重叠外，未观察到与任何视网膜特异性神经元标志物共表达Nestin。应该重新考虑巢蛋白作为一种独特的神经干细胞/祖细胞标记物的作用。在体外和体内，RPC成熟过程中Nestin的表达似乎不同。(C)2007爱思唯尔有限公司。保留所有权利。
The purpose of this study is to characterize the co-expression of nestin-a neuroectodermal stem cell and a reactive glial marker-with various mature retinal cell markers in retinal progenitor cells (RPCs) expanded in vitro, followed either by in vitro induction or subretinal transplantation. Rat RPCs derived from embryonic day (E) 17 rat retina were expanded in serum free defined culture, and induced to differentiate by alltrans retinoic acid (RA). Following induction, cells were stained for nestin in combination with retinal neuronal and glial markers. Cultured cells were collected for quantitative RT-PCR gene expression analysis prior to and after induction. In a second series, passage 2 RPCs were transplanted into the subretinal space of S334ter-3 retinal degeneration rats at postnatal day 28. After 1-4 weeks, sections through the transplant were double immunostained for nestin and various retinal specific neuronal markers. The cultured RPCs treated with RA exhibited nestin co-expression with various retinal specific markers, including protein kinase C alpha (PKC), neurofilament 200 (NTF200), cellular retinaldehyde binding protein (CRALBP), and rhodopsin. Following RA induction, quantitative RT-PCR analysis demonstrated downregulation of nestin, PAX-6, thy 1.1. and PKC alpha, and upregulation of rhodopsin, glial fibrillary acidic protein (GFAP), and CrX. No nestin coexpression was observed with any of the retinal specific neuronal markers in RPC transplants in vivo except for some nestin-immunoreactivity overlapping with GFAP positive cells in the host retina. The role of nestin as a unique neural stem/progenitor cell marker should be reconsidered. Nestin expression during RPC maturation appears to be different in vitro versus in vivo. (c) 2007 Elsevier Ltd. All rights reserved.