Claudin-3, an integral component of tight junction, has recently been shown to be expressed in retinal ganglion cells, retinal pigment cells, and retinal vascular endothelial cells. However, the role of claudin-3 in the development of the neural retina and its vessels remains undefined. This study aimed to investigate the role of zebrafish claudin-h (cldnh), the closest ortholog of mouse and human claudin-3, in the development of the neural retina and its vessels. Cldnh levels in green fluorescent protein transgenic zebrafish were genetically manipulated by cldnh morpholino oligonucleotide (MO) and cldnh mRNA to investigate gene function. The expression of cldnh was analyzed using polymerase chain reaction and immunofluorescence staining. The altered morphological, cellular and molecular events in the cldnh MO-morphant eyes were detected using hematoxylin-eosin staining, fluorescent dye injection, confocal in vivo imaging, BrdU labeling, TUNEL assay, RNA sequencing, and Western blot. We demonstrated that the cldnh protein was expressed in the neural retina and the hyaloid vessel which is the predecessor of the retinal vessel in zebrafish. Cldnh knockdown delayed lamination of the neural retina and reduced its thickness, which might be associated with the downregulation of the retinal development-related genes of atoh7, pcdh17, crx, neurod1, insm1a, sox9b and cdh11, and the upregulation of the cell cycle and apoptosis-associated genes of tp53, cdkn1a and casp8. Cldnh knockdown also reduced the density and interrupted the lumenization of the hyaloid vessels, which might be owing to the downregulation of the vessel formation-related genes of hlx1 and myl7. In conclusion, cldnh was required for the normal development of the neural retina and its vessels in zebrafish, providing a basis for elucidating its role in the pathogenesis of retinal vascular or inflammatory diseases.

Claudin-3是紧密连接的组成部分，最近已显示在视网膜神经节细胞，视网膜色素细胞和视网膜血管内皮细胞中表达。然而，claudin-3在神经视网膜及其血管发育中的作用仍然不确定。这项研究旨在调查斑马鱼claudin-h（cldnh），即小鼠和人类claudin-3的最接近直系同源物，在神经视网膜及其血管发育中的作用。cldnh吗啉代寡核苷酸（MO）和cldnh mRNA对绿色荧光蛋白转基因斑马鱼的cldnh水平进行了基因操作，以研究其基因功能。使用聚合酶链反应和免疫荧光染色分析cldnh的表达。形态改变了 使用苏木精-曙红染色，荧光染料注射，共聚焦体内成像，BrdU标记，TUNEL分析，RNA测序和Western印迹检测cldnh MO-morphant眼中的细胞和分子事件。我们证明了cldnh蛋白在斑马鱼的视网膜血管的前身-神经视网膜和玻璃状血管中表达。Cldnh击倒延迟了神经视网膜的层压并降低了其厚度，这可能与atoh7，pcdh17，crx，neurod1，insm1a，sox9b和cdh11的视网膜发育相关基因的下调以及细胞周期的上调和tp53，cdkn1a和casp8的凋亡相关基因。Cldnh的击倒也降低了密度，并中断了玻璃样血管的内腔流明，这可能是由于hlx1和myl7与血管形成相关的基因下调所致。总之，cldnh是斑马鱼神经视网膜及其血管正常发育所必需的，为阐明其在视网膜血管疾病或炎性疾病发病机理中的作用提供了基础。