Sánchez-Farías Nuria, Candal Eva
Department of Cell Biology and Ecology, University of Santiago de Compostela, 15782 Santiago de Compostela, Spain.
Exp Eye Res. 2015 May;134:90-100. doi: 10.1016/j.exer.2015.04.002. Epub 2015 Apr 4.
Doublecortin (DCX) is a microtubule-associated protein that has been considered a marker for neuronal precursors and young migrating neurons during the development of the central nervous system and in adult neurogenic niches. The retina of fishes represents an accessible, continuously growing and highly structured (layered) part of the central nervous system and, therefore, offers an exceptional model to extend our knowledge on the possible role of DCX in promoting neurogenesis and migration to appropriate layers. We have analyzed the distribution of DCX in the embryonic and postembryonic retina of a small shark, the lesser spotted dogfish Scyliorhinus canicula, by means of immunohistochemistry. We investigated the relationship between DCX expression and the neurogenic state of DCX-labeled cells by exploring its co-localization with the proliferation marker PCNA (proliferating cell nuclear antigen) and the marker of neuronal differentiation HuC/D. Since radially migrating neurons use radial glial fibers as substrate, we explored the possible correlation between DCX expression and cell migration along radial glia by comparing its expression with that of the glial marker GFAP (glial fibrillary acidic protein). Additionally, we characterized DCX-expressing cells by double immunocytochemistry using antibodies against Calbindin (a marker for mature bipolar and horizontal cells in this species) and Pax6, which has been proposed as a regulator of cell proliferation, cell differentiation, and neuron diversification in the neural retina of sharks. Strong DCX immunoreactivity was observed in immature cells and cell processes, at a time when retinal cells were not yet organized into different laminae. DCX was also found in subsets of mature ganglion, amacrine, bipolar and horizontal cells long after they had exited the cell cycle, a pattern that was maintained in juveniles and adults. Our results on DCX expression in the retina are compatible with a role for DCX in cell migration within the immature retina, and in dynamic neuronal plasticity in the mature retina. We also provide evidence of DCX expression in discrete cells in the retinal pigment epithelium of prehatching embryos and juveniles, which suggest that retinal pigmented epithelial cells in sharks, as in mammals, have an intrinsic capacity to proliferate and differentiate into cells with neural identity.
双皮质素（DCX）是一种微管相关蛋白，在中枢神经系统发育过程以及成体神经发生微环境中，一直被视为神经前体细胞和年轻迁移神经元的标志物。鱼类视网膜是中枢神经系统中一个易于研究、持续生长且结构高度有序（分层）的部分，因此，它为拓展我们对DCX在促进神经发生以及向合适层次迁移方面可能作用的认识提供了一个绝佳模型。我们通过免疫组织化学方法分析了小鲨鱼（斑点猫鲨Scyliorhinus canicula）胚胎期和胚后期视网膜中DCX的分布情况。我们通过探究DCX与增殖标志物增殖细胞核抗原（PCNA）以及神经元分化标志物HuC/D的共定位情况，研究了DCX表达与DCX标记细胞的神经发生状态之间的关系。由于径向迁移的神经元以放射状胶质纤维为底物，我们通过将DCX的表达与胶质标志物胶质纤维酸性蛋白（GFAP）的表达进行比较，探究了DCX表达与细胞沿放射状胶质细胞迁移之间的可能关联。此外，我们使用针对钙结合蛋白（该物种中成熟双极细胞和水平细胞的标志物）和Pax6的抗体，通过双重免疫细胞化学方法对表达DCX的细胞进行了表征，Pax6被认为是鲨鱼神经视网膜中细胞增殖、细胞分化和神经元多样化的调节因子。在视网膜细胞尚未组织成不同层时，在未成熟细胞和细胞突起中观察到了强烈的DCX免疫反应性。在成熟的神经节细胞、无长突细胞、双极细胞和水平细胞的亚群中，在它们退出细胞周期很久之后也发现了DCX，这种模式在幼体和成体中都持续存在。我们关于视网膜中DCX表达的结果与DCX在未成熟视网膜中的细胞迁移以及成熟视网膜中的动态神经元可塑性方面的作用相一致。我们还提供了在孵化前胚胎和幼体的视网膜色素上皮离散细胞中DCX表达的证据，这表明鲨鱼的视网膜色素上皮细胞与哺乳动物一样，具有增殖并分化为具有神经特性细胞的内在能力。
