Neural Tissue Engineering Based on Combinatorial Effect of Multiple Guidance Cues

基于多种引导线索组合效应的神经组织工程

• 批准号: 8674917
• 负责人: Mohammad Reza Abidian
• 金额: $ 30.22万
• 依托单位: PENNSYLVANIA STATE UNIVERSITY, THE
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-04-01 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8674917
• 关键词: Axon; Binding; Biochemical; Biocompatible; Caliber; Chemicals; Clinical; Complex; Cues; Development; Electrodes; Electroplating; Environment; Equilibrium; Growth; Growth Cones; Hand; Health; Hydrogels; In Vitro; Individual; Injury; Laminin; Left; Length; Measures; Microelectrodes; Modeling; Monitor; Morphology; Nanotubes; Natural regeneration; Nerve Growth Factors; Nerve Regeneration; Nervous system structure; Neurons; Pathway interactions; Peripheral Nervous System; Polymers; Proteins; Public Health; Rattus; Recovery of Function; Regulation; Research; Shapes; Side; Site; Societies; Solutions; Spinal Cord; Spinal Ganglia; Spinal cord injury; Surface; Techniques; Testing; Time; Tissue Engineering; Work; axon growth; axon guidance; axon regeneration; axonal guidance; base; combinatorial; controlled release; cost effective; density; electric impedance; extracellular; high throughput technology; improved; in vitro testing; in vivo; nanostructured; nerve gap; nerve injury; neurite growth; neuronal growth; novel; polymerization; public health relevance; relating to nervous system; research study; scaffold; statistical center; success; voltage

DESCRIPTION: This application aims to provide a mechanistic understanding of the effect of gradients of physical and chemical guidance cues (GCs), individually and combinatory, on guidance and modulation of axonal growth. The proposed study will specifically answer to questions whether: (1) immediate turning of growth cone depends on the difference between the concentration gradients on the left- and right-hand sides of the growth cone; (2) immediate and biased turning and growth-rate modulation work together to guide axons towards their targets; (3) integration of gradients of multiple cues can provide a precise regulation mechanism for axonal guidance. Axons are guided along specific pathways by gradients of attractive and repulsive cues in their extracellular environment. To understand the effect of gradients of guidance cues individually or in combination on growth cone turning and growth rate modulation, the development of platforms that are capable of producing precisely controlled shape gradients of guidance cues is essential. I propose to develop an inexpensive and high- throughput technology that is capable of providing precise, reproducible, and arbitrarily shaped gradients of physical and biochemical cues to direct and modulate axonal growth. For these studies, we will first fabricate aligned nanotubes of conducting polymer loaded with nerve growth factor on micro-fabricated electrode arrays. To release the entrapped nerve growth factor, we will actuate these nanotubes by applying electrical voltages. By varying the actuating voltage across the electrode array, we will create precisely controlled gradients of released nerve growth factor on these microelectrodes. Next, we will generate gradients of substrate-bound molecules, in this case laminin, on conducting polymer nanotubes across the electrode array. Inclusion of laminin on the nanotubes will be achieved by using this protein as a dopant during electropolymerization of conducting polymer. We will employ different concentrations of laminin on individual electrode sites to achieve the desired gradient profile. To generate gradients of surface topography, we will create gradients in diameter and surface roughness of aligned conducting polymer nanotubes on the micro-fabricated electrode arrays. We will modulate (a) the diameter of conducting polymer nanotubes by varying the time of electrochemical polymerization of conducting polymer, and (b) the surface roughness of conducting polymer nanotubes by varying the current density applied during electrodeposition. Finally, we will develop a 3D conduit consisting of a PDMS guidance channel that contains nanostructured conducting polymers that provide (i) physical and biochemical growth cues, and (ii) low impedance electrodes to monitor axonal growth by electrophysiological recording along the regeneration pathway. This multifunctional conduit will be tested in vitro and vivo to determine the effect of gradient of multiple guidance cues on axonal growth direction and rate. The results of these studies may significantly impact society by paving the way for a solution to the major clinical problem of axon regeneration and guidance.

描述：本应用程序旨在提供物理和化学引导信号（GCs）梯度（单独和组合）对轴突生长的引导和调节的机制理解。提出的研究将具体回答以下问题：(1)生长锥的立即转向是否取决于生长锥左右两侧浓度梯度的差异；(2)即时偏转与生长速率调节共同作用，引导轴突向目标方向运动；(3)多信号梯度的整合可为轴突定向提供精确的调控机制。轴突在细胞外环境中受到吸引和排斥信号的梯度引导，沿着特定的路径运动。为了了解单个或组合制导线索梯度对生长锥转向和生长速率调制的影响，开发能够产生精确控制制导线索形状梯度的平台至关重要。我建议开发一种廉价和高通量的技术，能够提供精确的、可重复的、任意形状的物理和生化信号梯度，以指导和调节轴突的生长。在这些研究中，我们将首先在微制造电极阵列上制备装载神经生长因子的导电聚合物排列纳米管。为了释放被困的神经生长因子，我们将通过施加电压来驱动这些纳米管。通过改变电极阵列上的驱动电压，我们将在这些微电极上创建精确控制的神经生长因子释放梯度。接下来，我们将产生底物结合分子的梯度，在这个例子中是层粘连蛋白，在电极阵列上导电聚合物纳米管。在导电聚合物的电聚合过程中，利用层粘连蛋白作为掺杂剂，可以将层粘连蛋白包裹在纳米管上。我们将在各个电极上使用不同浓度的层粘连蛋白来获得所需的梯度曲线。为了产生表面形貌的梯度，我们将在微制造电极阵列上产生排列的导电聚合物纳米管的直径和表面粗糙度梯度。我们将通过改变导电聚合物的电化学聚合时间来调节(a)导电聚合物纳米管的直径，以及(b)通过改变电沉积过程中施加的电流密度来调节导电聚合物纳米管的表面粗糙度。最后，我们将开发一种由PDMS引导通道组成的3D导管，该通道包含纳米结构的导电聚合物，可提供(i)物理和生化生长线索，以及（ii）低阻抗电极，通过电生理记录沿再生途径监测轴突生长。我们将在体外和体内测试这种多功能导管，以确定多种引导信号的梯度对轴突生长方向和速度的影响。这些研究结果通过为解决轴突再生和指导的主要临床问题铺平道路，可能会对社会产生重大影响。