Immobilizing Gradients of Neurotrophic Factors On An Aligned Biomaterial Scaffold

描述（由申请人提供）：用于促进脊髓损伤（SCI）后神经元再生和指导轴突生长的生物材料代表了治疗这种破坏性疾病的非常有前途的领域。特别地，模拟脊髓的高度对齐性质的材料将特别有效地引导轴突再生通过脊髓。 受伤的地方。许多研究表明，生长因子NGF（用于感觉神经元）和NT-3或BDNF（用于运动神经元）的梯度在刺激和指导SCI后轴突再生方面的有效性。所提出的工作概述了一种策略，用于用这些生长因子的梯度功能化高度对齐的生物材料水凝胶，以指导和 促进轴突再生。该生物材料将由肽两亲物（PA）分子组成，这些分子自组装成纳米纤维，并可用于形成可生物降解的水凝胶，其中纤维在宏观距离上对齐。申请人的初步工作已经证明，对齐的PAgel可以有效地捕获许多蛋白质数周，而不管大小和电荷如何，包括溶菌酶（其具有与大多数神经营养因子相同的大小和电荷）。此外，通过使用简单的基于扩散的方法，可以沿着对齐的轴构建蛋白质的稳定梯度，该方法允许在生物材料凝胶化之前立即直接产生梯度。在这项工作的基础上，将研究凝胶保留和产生稳定的生长因子梯度的能力。将详细研究凝胶在先前显示的促进生物活性的浓度范围内对神经营养因子的良好定义梯度的抑制能力。在构建凝胶后，将神经元掺入材料中以确定其促进和指导轴突生长的有效性。背根神经节神经元（其响应于NGF）和皮质脊髓运动神经元（其响应于NT-3）将被掺入具有适当神经营养因子梯度的PA凝胶中。轴突的长度和数量，以及它们的方向性和生长速率将被确定，并且梯度参数将被调整以优化轴突生长的这些测量。如果成功的话，这项工作将创造出第一种生物相容性，可降解和易于注射的生物材料，该材料在高度对齐和细胞粘附的支架上具有长寿命的生长因子梯度。从长远来看，这种材料可以在急性或慢性SCI后的损伤部位注射和凝胶化，以刺激轴突生长和功能连接，逆转或至少减轻损伤的影响。 公共卫生相关性：拟议的项目旨在合成一种高度对齐和可注射的支架，该支架具有长寿命的神经活性生长因子梯度，将为脊髓损伤中的神经元再生创造一种新型生物材料。固定在材料上的生长因子梯度将有助于刺激轴突生长，并将其引导到正确的方向。这种材料的长期应用将作为一种高度可转化的医学治疗，以帮助促进轴突再生，从而在脊髓损伤（急性或慢性）后重建功能连接，恢复感觉和运动控制并减轻损伤的有害影响。

DESCRIPTION (provided by applicant): Biomaterials for promoting neuron regeneration and directing axon growth following spinal cord injury (SCI) represent a highly promising field for treatment of this devastating condition. In particular, a material that mimics the highly aligned nature of the spinal cord will be particularly effective at directing axon regeneration through the site of injury. A number of studies have shown the effectiveness of gradients of growth factors NGF (for sensory neurons) and NT-3 or BDNF (for motor neurons), at both stimulating and directing axon regrowth following SCI. The proposed work outlines a strategy for functionalizing a highly aligned biomaterial hydrogel with gradients of these growth factors in order to direct and promote axon regeneration. The biomaterial will be composed of peptide amphiphile (PA) molecules that self-assemble into nanofibers and can be used to form a biodegradable hydrogel with the fibers aligned over macroscopic distances. Preliminary work by the applicant has demonstrated that aligned PAgels can efficiently entrap a number of proteins for several weeks, regardless of size and charge, including lysozyme (which has the same size and charge as most neurotrophins). In addition, a stable gradient of protein can be constructed along the aligned axis by using a simple diffusion-based method that allows for direct generation of the gradient immediately prior to biomaterial gelation. Building on this work, the ability of the gel t retain and create a stable gradient of growth factors will be investigated. The gel's ability to immobilize well-defined gradients of neurotrophins, in concentration ranges previously shown to promote bioactivity, will be investigated in detail. Following construction of the gel, neurons wil be incorporated into the material in order to determine its effectiveness at promoting and directing axon growth. Both dorsal root ganglia neurons (which respond to NGF), and corticospinal motor neurons (which respond to NT-3) will be incorporated into PA gels bearing the appropriate neurotrophin gradients. The length and number of axons, as well as their directionality and rate of growth will be determined, and the gradient parameters will be tuned to optimize these measures of axon growth. If successful, the work proposed will create the first biocompatible, degradable, and easily injectable biomaterial that possesses a long-lived gradient of growth factors on a highly aligned and cell-adhesive scaffold. In the long term, this material could be injected and gelled in the injury site following either acute or chronic SCI in order to stimulate axon growth and functional connectivity, reversing or at least mitigating the effects of the injury. PUBLIC HEALTH RELEVANCE: The proposed project, which seeks to synthesize a highly aligned and injectable scaffold bearing long-lived gradients of neuroactive growth factors, will create a novel biomaterial for neuron regeneration in spinal cord injury. The gradients of growth factors immobilized on the material will help stimulate axon growth, as well as direct it in the proper direction. The long term application of this material will be as a highly translatable medical treatment to help promote axon regeneration in order to reestablish functional connectivity following spinal cord injury (acute or chronic), restoring both sensory and motor control and mitigating the deleterious effects of the injury.