Photopolymerization-Induced Topography Directs Neurite and Schwann Cell Alignment

描述(由申请人提供)：与目标组织的不良整合严重限制了几乎所有神经假体的有效性。例如，由于神经-电极接口提供的空间和时间分辨率不佳，人工耳蜗(CI)接受者在复杂的听觉任务中表现不佳。引导螺旋神经节神经元(SGN)神经突起生长到与刺激电极紧密接近甚至接触的地方，可能会提高空间和时间分辨率，并提高性能。为了有用，再生必须是放射状的，并指向刺激的来源，重现正常的传入耳蜗神经。为了开发和理解引导SGN轴突和雪旺细胞(SC)生长的技术，利用光聚合(PP)技术，即用光形成聚合物，在生物相容的甲基丙烯酸酯聚合物中创建微通道。这些表面形态特征有力地指导了SGN轴突和SGSC的生长。我们假设，这些物理线索指导SGN突起和SGSC排列的能力取决于特定的表面形貌特征和材料特性，以及细胞内信号调节的结果，包括RhoA和Rho相关激酶(ROCK)。为了回应BRG PA10-009，本提案中的工作既是设计驱动的，也是假设驱动的。在目标1中，我们将利用PP提供的良好的空间反应控制来制作具有不同幅度、周期和表面纳米粗糙度的平行线空间光栅。这些地形特征对神经突起和神经胶质细胞黏附、存活和排列模式的影响程度将会被确定。由于细胞-材料的相互作用除了取决于表面形貌外，还取决于底物表面和机械性能，Aim 2决定了神经元和神经胶质细胞对不同表面(如极性)和机械(如硬度)属性的存活率和形态反应。最后，目标3检查了RhoA/ROCK，化学斥力线索引导神经突起的关键中介，对神经突起和SC向微图案排列的贡献。它还试图表征第二信使系统，包括环核苷酸、cAMP和cGMP，它们介导轴突和SC的排列以及RhoA/ROCK对表面形貌的反应。这些研究的结果将是第一批确定细胞感知和响应特定表面形貌和材料特性的基本机制之一。此外，他们将确定未来制造支架所需的地形特征和材料特性，这些支架可用于体内神经再生模型，包括设计增强型神经元：假体接口。

DESCRIPTION (provided by applicant): Poor integration with target tissue significantly limits the effectiveness of nearly all neural prostheses. For example, cochlear implant (CI) recipients perform poorly with complex auditory tasks due to poor spatial and temporal resolution provided by the neural-electrode interface. Directing growth of spiral ganglion neuron (SGN) neurites into close proximity, or even contact, with the stimulating electrodes would likely improve spatial and temporal resolution and increase performance. To be useful, regrowth must be radial and directed towards the source of stimulation, recapitulating normal afferent cochlear innervation. To develop and understand technology designed to guide SGN neurite and Schwann cell (SC) growth, photopolymerization (PP), i.e. the formation of polymers using light, was used to create microchannels in biocompatible methacrylate polymers. These surface topographic features robustly guide SGN neurite and SGSC growth. We hypothesize that the ability of these physical cues to direct SGN neurite and SGSC alignment depends on specific surface topographic features and material properties and results from tuning of intracellular signals including RhoA and Rho associated kinase (ROCK). In response to BRG PA10-009, the work in this proposal is both design- and hypothesis-driven. In aim 1 the excellent spatial reaction control afforded by PP will be leveraged to fabricate parallel line-space gratings with varied amplitude, periodicity, and surface nanoroughness. The extent to which these topographic features influence neurite and glial cell adhesion, survival and alignment to the pattern will then be determined. As cell-material interactions depend on substrate surface and mechanical properties in addition to surface topography, aim 2 determines the survival and morphological responses of neurons and glia to varied surface (e.g. polarity) and mechanical (e.g. stiffness) properties. Finally, aim 3 examines the contribution of RhoA/ROCK, key mediators of neurite guidance by chemorepulsive cues, to neurite and SC alignment to micropatterns. It also seeks to characterize second messenger systems including the cyclic nucleotides, cAMP and cGMP, that mediate neurite and SC alignment and RhoA/ROCK activity in response to surface topographies. The results of these studies will be among the first to define the basic mechanisms by which cells sense and respond to specific surface topographies and material properties. Furthermore, they will identify the topographic features and material properties necessary for future fabrication of scaffolds that can be used for in vivo neural regeneration models including the design of enhanced neuron:prosthesis interfaces.