Transgenic corneal neurofluorescence in mice: A new model for in vivo investigation of nerve structure and regeneration
Transgenic corneal neurofluorescence in mice: A new model for in vivo investigation of nerve structure and regeneration
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DOI：
10.1167/iovs.06-1192
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发表时间：
2007-04-01
影响因子：
4.4
通讯作者：
Rosenblatt, Mark I.
中科院分区：
文献类型：
--
作者：
Yu, Charles Q.;Rosenblatt, Mark I.
目的.目的：检测tby 1启动子调控下表达黄色荧光蛋白（YFP）的转基因小鼠角膜中神经元特异性荧光的水平，并探讨thy 1-YFP小鼠作为体内神经再生研究模型的可行性。用报告基因荧光技术观察了thy 1-YFP小鼠角膜神经支配的结构，并与传统免疫荧光技术观察的结构进行了比较。在全计数的角膜和三叉神经元培养物中测定具有YFP荧光的角膜神经的百分比。在体内监测创伤后荧光角膜神经元过程的再生。在小鼠角膜中，神经元特异性免疫染色确定神经以几束进入基质，然后延伸到整个角膜。这些基质神经束在角膜上皮下形成基底下神经丛。从这个神经丛的细神经旅行表浅到眼表面。神经元特异性表达的YFP允许可视化几乎所有的大神经束的基质，但只有一些许多更精细的神经的基底下神经丛和表面。在基底下神经丛中，46%的神经元突起显示AFP神经荧光。在体外培养的三叉神经元中，有22%的神经元表现出YFP神经荧光。角膜神经切断后，远侧的神经突起变性，而近侧的神经突起以不同于原始神经结构的方式再生。Thy 1-YFP小鼠显示神经荧光，并提供了一种新的模型，用于监测体内角膜神经的图案，损伤和生长。
PURPOSE. To quantify the level of neuron-specific fluorescence in the corneas of transgenic mice expressing yellow fluorescent protein (YFP) driven by the tby1 promoter and examine the viability of using thy1-YFP mice as a model for studying nerve regeneration in vivo.METHODS. The structure of corneal innervation in thy1-YFP mice visible with reporter gene fluorescence was compared with that Visible with traditional immunofluorescence techniques. The percentage of corneal nerves with YFP fluorescence in wholemounted corneas and trigeminal neuron cultures was determined. Regeneration of fluorescent corneal neuronal processes after wounding was monitored in vivo.RESULTS. In the mouse cornea, neuron-specific immunostaining determined that nerves enter the stroma in several bundles that then extend throughout the entire cornea. These stromal nerve bundles form a subbasal plexus beneath the corneal epithelium. Fine nerves from this plexus travel superficially to the ocular surface. Neuron-specific expression of YFP allowed visualization of nearly all large nerve bundles of the stroma but only some of the many finer nerves of the subbasal plexus and surface. In the subbasal nerve plexus, 46% of total neuronal processes exhibited AFP neurofluorescence. In vitro, 22% of cultured trigeminal neurons exhibited YFP neurofluorescence. After corneal nerve transection, nerve processes distal to the site of injury degenerated, whereas those proximal to the site regenerated in a pattern different from original nerve architecture.CONCLUSIONS. Thy1-YFP mice display neurofluorescence and provide a novel model for monitoring the patterning, injury, and growth of corneal nerves in vivo.