TISSUE ENGINEERED CELL TRANSPLANTATION FOR GLAUCOMA AND OPTIC NEUROPATHIES

项目总结/摘要 在这里，我们建议开发一种生物工程支架，用于内层视网膜， 既可以作为这种组织的模型，也可以作为细胞移植的工具。神经视网膜，就像视网膜的其他部分一样， 中枢神经系统（CNS）在与损伤或疾病相关的细胞死亡后不能再生。到 为了克服这种再生修复的缺乏，在光感受器的疾病中，细胞替代疗法已经 被尝试。在这些研究中，注射的细胞能够迁移到视网膜的正确层中， 突触和功能的动物模型。1，2但这样的技术可以用于视网膜疾病， 神经节细胞（RGC），其轴突必须通过患病视神经的抑制环境再生 神经？最近发现的促进视神经和CNS轴突再生的分子机制， 例如PTEN/SOCS 3或KLF 4缺失，3-6结合我们的研究显示移植的RGC延长 局部突起并在内丛状层（IPL）形成突触，7表明移植治疗 对RGC来说是可能的。然而，迄今为止的数据表明，移植的细胞在很大程度上不能 将轴突导向视神经头，这可能是由于视网膜引导的发育变化 最近，我们开发了一种可生物降解的径向静电纺丝支架（rES）细胞递送系统， 能够放射状引导RGC轴突、匹配视网膜神经纤维层方向的载体 然而，接种在rES上的RGC在两个方向上生长，而不是使其轴突生长。 像在天然组织中一样向中心极化。此外，尚不清楚移植的细胞是否会 能够将它们的树突从细胞递送载体上延伸出来，与受损的视网膜形成突触。 在这项研究中，我们将进一步发展我们的细胞输送支架，固定在细胞生长过程中发现的神经营养因子。 一旦移植，神经轴突向支架中心和视神经乳头生长。 此外，我们将联合收割机与由ECM基质成分组成的水凝胶结合， IPL以刺激移植的RGC树突向视网膜植块延伸。使用这些外植体 模型，我们将评估移植细胞的功能性突触连接通过染色和 光响应的传播。最后，使用经过改造的细胞通过受损的视神经再生，我们将 研究移植细胞的轴突进入视神经的能力。这些研究将导致 具有神经节细胞层和水凝胶的正确图案化的视网膜细胞递送装置的产生 系统优化以刺激移植细胞的整合。总的来说，这一建议将是一个 我们长远目标是帮助青光眼及其他视神经疾病患者恢复视力， 神经病

PROJECT SUMMARY/ABSTRACT Here we propose to develop a bio-engineered scaffold for the inner retina capable of being used as both a model for this tissue and as a tool for cell transplantation. The neural retina, like other parts of the central nervous system (CNS), fails to regenerate following cell death associated with injury or diseases. To overcome this lack of regenerative repair, in diseases of the photoreceptors, cell replacement therapies have been attempted. In these studies, injected cells are able to migrate into the correct lamina of the retina, form synapses and function in animal models.1, 2 But can such a technique be used with diseases of the retinal ganglion cells (RGCs) whose axons must regrow through the inhibitory environment of the diseased optic nerve? The recent discovery of molecular mechanisms that promote optic nerve and CNS axon regeneration, such as PTEN/SOCS3 or KLF4 deletion,3-6 combined with our studies showing transplanted RGCs extend processes locally and form synapses in the inner plexiform layer (IPL),7 suggest that a transplantation therapy may yet be possible for RGCs. However, data thus far suggest that transplanted cells are largely unable to direct their axons towards the optic nerve head, perhaps due to developmental changes in retinal guidance molecules.8, 9 Recently, we have developed a biodegradable radial electrospun scaffold (rES) cell delivery vehicle capable of directing RGC axons radially, matching the orientation of the retinal nerve fiber layer (NFL).10 However the RGCs seeded on the rES grow in both directions rather than having their axon growth polarized towards the center as it is in the native tissue. In addition, it is not known if the transplanted cells will be capable of extending their dendrites off of the cell delivery vehicle to form synapses with the injured retina. In this study, we will further develop our cell delivery scaffold, immobilizing neurotrophic factors found during development to polarize axon growth towards the scaffold center and the optic nerve head once transplanted. In addition, we will combine the rES with a hydrogel composed of ECM matrix components of the developing IPL in order to stimulate the transplanted RGC dendrites extension to retinal explants. Using these explant models, we will evaluate the transplanted cells for functional synaptic connections through staining and the propagation of light responses. Finally, using cells modified for regrowth through an injured optic nerve, we will investigate the ability of axons from transplanted cells to enter the optic nerve. These studies will lead to the creation of a retinal cell delivery device with the correct patterning of the ganglion cell layer and a hydrogel system optimized to stimulate the integration of the transplanted cells. Taken together, this proposal will be an important step towards our long term goal of restoring vision to those suffering from glaucoma and other optic neuropathies.