Ruiz Department of Ophthalmology & Visual Science, McGovern Medical School, The University of Texas Health Science Center at Houston, Houston, TX 77030, USA.
The MD Anderson Cancer Center/UTHealth Graduate School of Biomedical Sciences, Houston, TX 77030, USA.
Cells. 2020 Oct 6;9(10):2242. doi: 10.3390/cells9102242.
More than 1.5 million people suffer from Retinitis Pigmentosa, with many experiencing partial to complete vision loss. Regenerative therapies offer some hope, but their development is challenged by the limited regenerative capacity of mammalian model systems. As a step toward investigating regenerative therapies, we developed a zebrafish model of Retinitis Pigmentosa that displays ongoing regeneration. We used Tol2 transgenesis to express mouse rhodopsin carrying the P23H mutation and an epitope tag in zebrafish rod photoreceptors. Adult and juvenile fish were examined by immunofluorescence, TUNEL and BrdU incorporation assays. P23H transgenic fish expressed the transgene in rods from 3 days post fertilization onward. Rods expressing the mutant rhodopsin formed very small or no outer segments and the mutant protein was delocalized over the entire cell. Adult fish displayed thinning of the outer nuclear layer (ONL) and loss of rod outer segments, but retained a single, sparse row of rods. Adult fish displayed ongoing apoptotic cell death in the ONL and an abundance of proliferating cells, predominantly in the ONL. There was a modest remodeling of bipolar and Müller glial cells. This transgenic fish will provide a useful model system to study rod photoreceptor regeneration and integration.
超过 150 万人患有色素性视网膜炎，许多人经历部分至完全视力丧失。再生疗法带来了一些希望，但由于哺乳动物模型系统的再生能力有限，其发展受到了挑战。作为研究再生疗法的一个步骤，我们开发了一种具有持续再生能力的色素性视网膜炎斑马鱼模型。我们使用 Tol2 转座酶在斑马鱼杆状光感受器中表达携带 P23H 突变和表位标签的小鼠视紫红质。通过免疫荧光、TUNEL 和 BrdU 掺入测定法检查成年鱼和幼鱼。P23H 转基因鱼从受精后 3 天开始在杆状细胞中表达转基因。表达突变视紫红质的杆状细胞形成非常小或没有外节，并且突变蛋白在整个细胞中弥散分布。成年鱼表现出外核层 (ONL) 的变薄和杆状细胞外节的丧失，但保留了单一、稀疏的杆状细胞列。成年鱼的 ONL 中持续发生凋亡性细胞死亡，并且有大量增殖细胞，主要位于 ONL 中。双极细胞和 Müller 胶质细胞有适度的重塑。这种转基因鱼将为研究杆状光感受器再生和整合提供一个有用的模型系统。
