Manipulating retinal progenitor cells

描述(申请人提供)：本次修订的R21申请的总体目标是确定小鼠视网膜前体细胞(RPC)在其正常胚胎环境之外分化为视网膜神经节细胞(RGC)的潜力。胚胎或胚胎干细胞来源的RPC在成人视网膜中分化为功能性RGCs的潜力将通过将它们移植到RGCs已被基因去除的成人宿主视网膜中在体内进行检测。一种新的遗传性小鼠模型已经被开发出来，在该模型中，部分或全部视网膜节细胞在成年后的任何时候都可以被消融，从而导致视神经退化。这些小鼠的视网膜为细胞移植提供了独特的微环境，以恢复视网膜节细胞并再生视神经或防止其进一步退化。实验计划依赖于理解在视网膜形成过程中控制RGC规范和分化的关键调控事件。原神经性基本螺旋-环-螺旋因子Math5在控制RGC发育的基因调控网络中占有中心节点，因为它负责赋予RPC获得RGC命运的能力。需要检验的假设是，表达Math5的RPC是一个独特的、可分离的RPC亚群，即使将Math5表达的RPC放置在成人视网膜的微环境中，也将保持其发育潜力。为了研究表达Math5的RPC与成人视网膜微环境的相互作用，我们将研究纯化的表达Math5的RPC在移植到RGC切除的成人视网膜后分化为功能性RGC的可能性。确定胚胎RPC适应成人视网膜微环境的发育潜力将有助于理解视网膜的发育，并建立更强大的方法来预防、修复和再生受损的视网膜。成年小鼠视网膜神经节细胞的基因切割为视网膜再生提供了一种新的模型。了解RPC在正常环境中分化为RGC的机制，为从胚胎视网膜或多能胚胎干细胞生成大量具有功能的RGC前体细胞奠定了基础。 与公共卫生相关：该项目将揭示胚胎视网膜前体细胞的特性，这些细胞在成人神经视网膜中形成视网膜神经节细胞。实验旨在确定视网膜前体细胞适应成人视网膜微环境的潜力，而不是适应正常的胚胎微环境。该项目将有助于了解视网膜的发育，并建立新的方法来再生受损的视网膜，防止或修复视神经炎、缺血性视神经病变和青光眼等视网膜疾病的视神经退化。

DESCRIPTION (provided by applicant): The overall objective of this revised R21 application is to determine the potential of mouse retinal progenitor cells (RPCs) to differentiate into retinal ganglion cells (RGCs) outside of their normal embryonic environment. The potential of embryo- or embryonic stem cell-derived RPCs to differentiate into functional RGCs in adult retinas will be examined in vivo by transplanting them into adult host retinas in which RGCs have been genetically ablated. A novel genetic mouse model has been developed in which some or all RGCs can be ablated at any time during adult life with the consequent degeneration of the optic nerve. Retinas from these mice provide a unique microenvironment for cell transplantation approaches to restore RGCs and regenerate the optic nerve or prevent its further degeneration. The experimental plan depends on understanding the key regulatory events that control the specification and differentiation of RGCs during retinogenesis. The proneural basic helix-loop-helix factor Math5 occupies a central node in the gene regulatory network that controls RGC development because it is responsible for endowing RPCs with the competence to acquire a RGC fate. The hypothesis to be tested is that Math5-expressing RPCs are a distinct, isolatable RPC subpopulation and that Math5-expressing RPCs will retain their developmental potential even when placed into the microenvironment of the adult retina. To investigate the interactions between Math5-expressing RPCs and the adult retinal microenvironment, the potential of purified Math5-expressing RPCs to differentiate into functional RGCs after transplantation into RGC- ablated adult retinas will be investigated. Determining the developmental potential of embryonic RPCs to adapt to the microenvironment of the adult retina will contribute towards an understanding of retina development as well as establishing more robust methods to prevent, repair and regenerate damaged retinas. Genetic ablation of RGCs in adult mice provides a new model for retinal regeneration. Knowledge of the mechanisms that allow RPCs to differentiate into RGCs in their normal environment provides the underpinning for generating large numbers of functional RGC progenitors from embryonic retinas or pluripotent embryonic stem cells. PUBLIC HEALTH RELEVANCE: This project will reveal new insights into the properties of the embryonic retinal progenitor cells that give rise to retinal ganglion cells in the adult neural retina. Experiments are designed to determine the potential of retinal progenitor cells to adapt to the microenvironment of the adult retina rather than their normal embryonic microenvironment. The project will contribute to an understanding of retinal development as well as establishing new approaches to regenerate damaged retinas and prevent or repair optic nerve degeneration in retinal pathologies such as optic neuritis, ischemic optic neuropathy, and glaucoma.