A nano-enabled biomimetic platform for neuronal differentiation and maturation

项目摘要 获得成熟的细胞和组织是了解疾病的病理生物学和发育的关键 药物疗法。神经细胞的细胞和分子研究是现代神经科学的基石，但 大细胞神经元(MCN)和某些类型的干细胞违背了传统的培养方法。20多年来， 体外培养MCNs的挑战一直是细胞和分子生物学研究的障碍。 催产素和加压素激素、神经递质及其受体的产生和调节。为 原因不明，MCN在没有饲养层的情况下不能在体外生长。传统的基板还没有 能够取代这些相互依赖的需求。这项提议的目标是利用身体接触 和细胞外基质的化学特征，以创建可靠地促进， 在体外维持和成熟MCN。我们描述了一个两阶段的方法，它最大限度地提高了进度 在推进创新的同时管理风险，以解决MCN文化挑战。首先，我们将测试和 脑脱细胞外基质(Brain-DECM)与常规方法改善MCN的比较 产量超过了已建立的共同培养模式。这将大大减少时间、可变性和 通过改进细胞培养过程来取代饲养层，减少动物使用，从而使实验变得复杂。 由于脑-DECM是一种具有许多未知数的复杂试剂，我们的第二阶段是创建纳米级 仿生神经胶质细胞外基质。其结果是一种组织芯片，它保留了生长许可 物理提示，同时结合来自神经胶质细胞的有助于发育的生化配体。这块组织 芯片平台可能会提高神经元产量并增加获取成熟MCNs的途径，以实现高分辨率成像 以及对难以培养的细胞群体的分析。我们希望这一创新的解决方案能够长期- 长期的挑战将是可靠的、多产的和有影响力的，以产生新的药物发现机会 翻译科学和医学。

Project Summary Access to mature cells and tissues is key to understanding the pathobiology of diseases and for developing drug therapies. Cellular and molecular studies of nerve cells are a cornerstone of modern neuroscience, yet magnocellular neurons (MCN) and some stem cell types defy conventional culture methods. For over 20 years, the challenge of culturing MCNs in vitro has been an obstacle for cellular and molecular studies on the production and regulation of oxytocin and vasopressin hormones, neurotransmitters, and their receptors. For unknown reasons, MCN cannot be grown in vitro without feeder layers. Conventional substrates have not yet been able to replace these co-dependent requirements. The goal of this proposal is to harness physical contacts and chemical signatures of the extracellular matrix to create a cell culture platform that reliably promotes, maintains, and matures MCN in vitro. We describe a two-stage approach that maximizes the progress and manages the risks while advancing innovation to solve the MCN culture challenge. First, we will test and compare brain decellularized extracellular matrix (Brain-DECM) to conventional methods to improve MCN yields beyond the established co-culture model. This will substantially reduce the time, variability, and complexity of experiments by refining the cell culture process to replace feeder-layers and decrease animal use. Because Brain-DECM is complex reagent with many unknowns, our second stage is to create a nano-scale biomimetic glial-derived extracellular matrix. The result is a tissue chip that retains the growth-permissive physical cues while incorporating developmentally-instructive biochemical ligands derived from glia. This tissue chip platform may improve neuron yield and increase access to mature MCNs to enable high-resolution imaging and analyses for difficult to culture cellular populations. We expect that this innovative solution to a long- standing challenge will be reliable, productive, and impactful to yield new opportunities for drug discovery for translational science and medicine.