Generation of Retinal Ganglion-like Cells from Reprogrammed Mouse Fibroblasts
Generation of Retinal Ganglion-like Cells from Reprogrammed Mouse Fibroblasts
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DOI：
10.1167/iovs.09-4504
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发表时间：
2010-11-01
影响因子：
4.4
通讯作者：
Ge, Jian
中科院分区：
文献类型：
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作者：
Chen, Mengfei;Chen, Qin;Ge, Jian
目的.体细胞可以通过Oct-3/4、Sox 2、c-Myc和Klf 4重编程为胚胎干细胞样多能状态。Sox 2作为一种重要的重编程因子，也有助于眼睛和视网膜的发育。本研究旨在探讨诱导性多能干细胞（iPS）是否表达视网膜祖细胞（RPC）相关基因，以及iPS细胞能否直接分化为视网膜神经节细胞（RGCs）。用小鼠尾尖成纤维细胞（TTFs）异位表达的4种因子诱导小鼠iPS细胞。采用RT-PCR和免疫荧光法检测iPS细胞中RPC相关基因的表达。通过添加Dkk 1 + Noggin（DN）+ DAPT和过表达Math 5来诱导iPS细胞分化成RGCs。将iPS衍生的视网膜神经节（RG）样细胞注射到视网膜中，并通过荧光化学分析眼睛。iPS细胞固有地表达RPC相关基因，例如Pax 6、Rx、Otx 2、Lhx 2和Nestin。过表达Math 5和加入DN可使iPS直接分化为视网膜神经节样细胞。这些iPS衍生的RG样细胞显示长突触和基因表达模式，包括Math 5，Brn 3b，Islet-1和Thy1.2。此外，通过DAPT抑制Hes 1会增加RGC标记基因的表达。此外，iPS诱导的视网膜节细胞样细胞移植后能够存活，但不能整合到正常视网膜中。四因子iPS细胞固有地表达RPC相关基因，并且iPS细胞可通过转录因子表达的调控进一步向RG样细胞转化。这些发现表明iPS细胞对于视网膜变性疾病的再生研究是有价值的。（Invest Ophthalmol维斯科学。2010;51：5970-5978）DOI：10.1167/iovs.09-4504
PURPOSE. Somatic cells can be reprogrammed into an embryonic stem cell-like pluripotent state by Oct-3/4, Sox2, c-Myc, and Klf4. Sox2 as an essential reprogramming factor also contributes to the development of the eye and the retina. This study was conducted to determine whether induced pluripotent stem (iPS) cells express retinal progenitor cell (RPC)-related genes and whether iPS cells can directly differentiate into retinal ganglion cells (RGCs).METHODS. Mouse iPS cells were induced by the ectopically expressed four factors in tail-tip fibroblasts (TTFs). The expression of RPC-related genes in iPS cells was analyzed by RT-PCR and immunofluorescence. iPS cells were induced to differentiate into RGCs by the addition of Dkk1 + Noggin (DN) + DAPT and overexpression of Math5. iPS-derived retinal ganglion (RG)-like cells were injected into the retina, and the eyes were analyzed by immunohistochemistry.RESULTS. iPS cells inherently express RPC-related genes such as Pax6, Rx, Otx2, Lhx2, and Nestin. Overexpression of Math5 and addition of DN can directly differentiate iPS into retinal ganglion-like cells. These iPS-derived RG-like cells display long synapses and gene expression patterns, including Math5, Brn3b, Islet-1, and Thy1.2. Furthermore, inhibiting Hes1 by DAPT increases the expression of RGC marker genes. In addition, iPS-derived RG-like cells were able to survive but were unable to be integrated into the normal retina after transplantation.CONCLUSIONS. The four factor iPS cell inherently expressed RPC-related genes, and the iPS cell could be further turned into RG-like cells by the regulation of transcription factor expression. These findings demonstrate that iPS cells are valuable for regeneration research into retinal degeneration diseases. (Invest Ophthalmol Vis Sci. 2010;51:5970-5978) DOI:10.1167/iovs.09-4504