Photopolymerization-Induced Topography Directs Neurite and Schwann Cell Alignment

描述（由申请人提供）：与目标组织的整合不良严重限制了几乎所有神经假体的有效性。例如，由于神经电极界面提供的空间和时间分辨率较差，人工耳蜗（CI）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）神经突的生长与刺激电极接近甚至接触，可能会提高空间和时间分辨率并提高性能。为了有效，再生必须是径向的，并指向刺激源，再现正常的传入耳蜗神经支配。为了开发和理解引导SGN神经突和雪旺细胞（SC）生长的技术，光聚合（PP），即利用光形成聚合物，被用于在生物相容性甲基丙烯酸酯聚合物中创建微通道。这些表面地形特征有力地指导了SGN神经突和SGSC的生长。我们假设这些物理线索指导SGN神经突和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（包括RhoA和Rho相关激酶（ROCK））调谐的结果。作为对BRG PA10-009的回应，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，PP提供的出色的空间反应控制将被用来制造具有不同振幅、周期性和表面纳米粗糙度的平行线空间光栅。这些地形特征在多大程度上影响神经突和神经胶质细胞的粘附、存活和对齐模式，然后将确定。由于细胞-物质相互作用除了依赖于表面形貌外，还依赖于基质的表面和机械特性，因此目标2决定了神经元和胶质细胞对不同表面（如极性）和机械特性（如刚度）的存活和形态反应。最后，目的3研究了RhoA/ROCK（通过化学排斥信号引导神经突的关键介质）对神经突和SC对齐微模式的贡献。它还试图表征第二信使系统，包括环核苷酸、cAMP和cGMP，它们介导神经突起和SC排列以及RhoA/ROCK活性，以响应表面地形。这些研究的结果将是第一个定义细胞感知和响应特定表面地形和材料特性的基本机制。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：假体界面。

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.