Combinatorial Manipulation of Transcription Factors to Promote CNS Regeneration

项目摘要 在成人中，中枢神经系统（CNS）中的轴突在因损伤或损伤而丧失后通常不能再生。 疾病，导致永久性和不可治愈的残疾。轴突生长受到不利生长环境的阻碍， 以及随着CNS神经元老化轴突生长的内在能力的发育丧失。转录 转录因子（TF）与DNA相互作用，并协调广泛的细胞物质的产生， 成为重要的治疗靶点，以提高受损神经元的再生能力。例如，强迫 在成年神经元中重新表达一种名为KLF 6的促再生TF可以提高其轴突生长的能力 脊髓损伤后。我们现在将测试三个相辅相成的战略，以加强 KLF 6的有前途的促再生特性。首先，使用新的生物信息学管道，我们预测 与KLF 6功能性相互作用的其他TF，并证实了它们协同增强轴突生长的能力 在轴突生长的细胞培养模型中与KLF 6组合。因此，我们将进行三项体内测试 选择的因素，EOMES，NR 5A 2和RARB，用于增强KLF 6的促再生特性的能力， 脊髓损伤的动物模型。第二，我们将通过移植生长来补充这些TF干预措施- 移植到脊髓损伤的部位。这些移植物将缓解脊髓中持续的生长抑制 脐带环境，从而揭露TF治疗的促再生作用。最后，我们将利用一个新的 开发的基因治疗载体，能够以前所未有的效率逆行传递基因。注射 这种载体的脊髓导致广泛的基因表达在整个受损神经元 脑干中脑和运动皮层这种输送系统涉及更多数量和更广泛的多样性， 细胞类型比目前的直接脑注射的做法，从而最大限度地实现功能的机会， 脊髓损伤后的增益。在这些目标中，组织清除和3D显微镜将揭示新的解剖结构， 诱发再生的细节将这些尖端改进汇集到以TF为中心的战略中 将使该领域朝着新颖有效的治疗方法发展， CNS损伤的后果。

PROJECT SUMMARY In adults, axons in the central nervous system (CNS) generally fail to regenerate after they are lost to injury or disease, leading to permanent and incurable disability. Axon growth is prevented by a hostile growth environment, as well as a developmental loss in the intrinsic capacity for axon growth as CNS neurons age. Transcription factors (TFs) interact with DNA and coordinate the production of broad sets of cellular materials, and have emerged as important therapeutic targets to boost regenerative ability within injured neurons. For example, forced re-expression of a pro-regenerative TF called KLF6 in adult neurons can improve their capacity for axon growth after spinal injury. We will now test three complementary and mutually supportive strategies to enhance the promising pro-regenerative properties of KLF6. First, using a novel bioinformatics pipeline, we have predicted additional TFs that functionally interact with KLF6 and verified their ability to synergistically enhance axon growth when combined with KLF6 in cell culture models of axon growth. We will therefore perform in vivo tests of three selected factors, EOMES, NR5A2, and RARB, for the ability to enhance KLF6’s pro-regenerative properties in animal models of spinal cord injury. Second, we will supplement these TF interventions with transplants of growth- permissive stem cells into sites of spinal injury. These grafts will alleviate persistent growth inhibition in the spinal cord environment, and thus unmask the pro-regenerative effects of TF treatments. Finally, we will harness a newly developed gene therapy vector that enables retrograde delivery of genes with unprecedented efficiency. Injection of this vector to the spinal cord results in widespread gene expression in injured neurons throughout the brainstem, midbrain, and motor cortex. This delivery system engages a larger number and a wider diversity of cell types than the current practice of direct brain injection, thus maximizing the chance of achieving functional gains after spinal injury. Throughout these aims, tissue clearing and 3D microscopy will reveal new anatomical details of the evoked regeneration. Bringing together these cutting-edge improvements to a TF-centered strategy will move the field toward novel and effective treatments for individuals suffering from the debilitating consequences of CNS injury.