Pax6- and Six3-mediated induction of lens cell fate in mouse and human ES cells.
Pax6- and Six3-mediated induction of lens cell fate in mouse and human ES cells.
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DOI：
10.1371/journal.pone.0115106
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Maas RL
中科院分区：
文献类型：
--
作者：
Anchan RM;Lachke SA;Gerami-Naini B;Lindsey J;Ng N;Naber C;Nickerson M;Cavallesco R;Rowan S;Eaton JL;Xi Q;Maas RL
胚胎干细胞（ES）为体内组织分化的研究提供了一个潜在的有用的体外模型。我们利用小鼠和人胚胎干细胞研究晶状体调控基因Pax6和Six3是否能在体外诱导晶状体细胞的命运。为了帮助评估晶状体分化的开始，我们获得了一种新的mES细胞系（Pax6-GFP mES），该细胞系在Pax6 P0启动子和晶状体外胚层增强子的控制下表达GFP报告基因。通过转染或慢病毒感染将Pax6或Six3表达载体导入mES或hES细胞，分析分化的ES细胞中晶状体标志物的表达情况。转染Pax6或Six3而不转染其他基因的mES细胞可诱导晶状体细胞标记物的表达，并在转染3天后上调Pax6-GFP mES细胞中GFP报告基因的表达。转染后7天，mES细胞培养中表达γ a -晶体蛋白（一种晶状体纤维细胞分化标志物）的菌落数量比对照组增加了10倍。RT-PCR和免疫染色显示诱导了额外的晶状体上皮或纤维细胞分化标志物，包括Foxe3、Prox1、α-和β-晶体蛋白以及Tdrd7。在培养30 d后，可形成表达γ - a -crystallin或prox1的透镜状体。在hES细胞中，Pax6或Six3慢病毒载体也诱导晶状体标志物表达。表达晶状体标记的胚胎干细胞与Pax6或Six3转导的细胞接近但不同，这表明后者通过非细胞自主机制诱导附近未分化的胚胎干细胞采用晶状体命运。总之，我们描述了一种新的胚胎干细胞GFP报告系，可用于监测晶状体命运的诱导，并证明Pax6或Six3足以诱导胚胎干细胞采用晶状体命运，可能通过非细胞自主机制。这些发现将有助于晶状体发育的研究。
Embryonic stem (ES) cells provide a potentially useful in vitro model for the study of in vivo tissue differentiation. We used mouse and human ES cells to investigate whether the lens regulatory genes Pax6 and Six3 could induce lens cell fate in vitro. To help assess the onset of lens differentiation, we derived a new mES cell line (Pax6-GFP mES) that expresses a GFP reporter under the control of the Pax6 P0 promoter and lens ectoderm enhancer. Pax6 or Six3 expression vectors were introduced into mES or hES cells by transfection or lentiviral infection and the differentiating ES cells analyzed for lens marker expression. Transfection of mES cells with Pax6 or Six3 but not with other genes induced the expression of lens cell markers and up-regulated GFP reporter expression in Pax6-GFP mES cells by 3 days post-transfection. By 7 days post-transfection, mES cell cultures exhibited a>10-fold increase over controls in the number of colonies expressing γA-crystallin, a lens fiber cell differentiation marker. RT-PCR and immunostaining revealed induction of additional lens epithelial or fiber cell differentiation markers including Foxe3, Prox1, α- and β-crystallins, and Tdrd7. Moreover, γA-crystallin- or Prox1-expressing lentoid bodies formed by 30 days in culture. In hES cells, Pax6 or Six3 lentiviral vectors also induced lens marker expression. mES cells that express lens markers reside close to but are distinct from the Pax6 or Six3 transduced cells, suggesting that the latter induce nearby undifferentiated ES cells to adopt a lens fate by non-cell autonomous mechanisms. In sum, we describe a novel mES cell GFP reporter line that is useful for monitoring induction of lens fate, and demonstrate that Pax6 or Six3 is sufficient to induce ES cells to adopt a lens fate, potentially via non-cell autonomous mechanisms. These findings should facilitate investigations of lens development.
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影响因子：
64.8
作者：
HILL, RE;FAVOR, J;VANHEYNINGEN, V
通讯作者：
VANHEYNINGEN, V
影响因子：
30.8
作者：
JORDAN, T;HANSON, I;VANHEYNINGEN, V
通讯作者：
VANHEYNINGEN, V
DOI：
10.1073/pnas.132195699
发表时间：
2002-06-25
影响因子：
11.1
作者：
Goudreau, G;Petrou, P;Gruss, P
通讯作者：
Gruss, P
影响因子：
10.5
作者：
Ashery-Padan, R;Marquardt, T;Gruss, P
通讯作者：
Gruss, P
影响因子：
11.4
作者：
Liu, Wei;Lagutin, Oleg V.;Oliver, Guillermo
通讯作者：
Oliver, Guillermo