Computational and Cell Culture Models for Mucus Clearance

描述(由申请人提供)：本申请涉及广泛的挑战领域(06)使能技术和具体挑战主题06-ES-102*：3-D或虚拟模型以减少研究中动物的使用：创建微型多细胞器官，用于高通量筛选化学毒性测试。随着器官系统功能计算模型的发展，复制主要器官系统的宏观结构和功能的多种细胞类型的新型微观系统的开发，可以加快对环境中多种化学物质的毒性测试。进一步产生3-D生物模型的研究将为快速筛选肺和肝脏等器官的毒性提供新的分析方法。在这些系统中使用的细胞类型，如人类干细胞，将减少对动物的使用，并改善我们对环境中化学危害的评估。联系人：David Balshaw博士，Balshaw@niehs.nih.gov，(919)541-2448标题：粘液清除的计算和细胞培养模型摘要：肺部每天吸入100多万种感染性和有毒物质。对肺的防御始于排列在上皮细胞层的粘液层。当这些攻击到达粘液/空气界面时，感染和清除之间的竞赛就开始了。微粒、病原体和过敏原在细胞膜上扩散，然后才能通过粘液的携带流动被清除。为了提供有效的许可，该机构必须实现几个目标。首先，它必须为粘液屏障层提供高度和粘弹性，从而为感染性病原体提供显著的传播时间。其次，通过纤毛或气流的推进机制必须使粘液以一定的速度移动，以便在它们扩散到细胞膜之前清除这些制剂。第三，粘液的粘弹性必须足以防止呼吸道泛滥，但又必须具有足够的流动性，以允许纤毛和气流进行运输。该项目的目标是生成基于计算和细胞培养的粘液清除模型。由于清除失败可以理解为一系列肺衰竭情景的第一步，因此建立有效的毒性测试模型系统至关重要。基于细胞培养模型的使用允许包含复杂的生化和免疫反应。纤毛培养的使用已经被证明可以产生和维持适当的粘液层，并产生用于推进的协调排列的纤毛，这是我们模型系统的起点。我们将产生第一个纤毛细胞培养系统，将细胞置于复制肺基本特征的几何形状中：具有收敛横截面的定向、线性流动，可挑战垂直运输以对抗重力。这个模型系统将有可能作为环境效应器的敏感AS-比方说，妥协粘液高度调节，粘液流变学，纤毛密度和协调。我们将在微流控系统中进一步开发该分析方法，该系统有12个独立的分析方法并行运行。这将使这一复杂的基于细胞的分析方法进入中通量筛选。除了生物物理模型，我们还需要一个计算模型，以便我们可以预测环境攻击的后果并设计有效的治疗方法。我们将开发理论模型，研究纤毛和气流的推进机制。后者在有效的清除动作中起作用，如咳嗽，可以理解的是，即使在没有纤毛推进的情况下，咳嗽也可以维持无菌的呼吸道。然而，目前还没有粘液流变学和层厚、纤毛有效性和气流在产生足够的清除以维持健康的肺功能方面的作用的预测模型。通过结合来自应用数学、物理、生物化学和生物物理学的研究小组，以及北卡罗来纳大学囊性纤维化中心，该项目的目标是开发基于细胞的生物物理分析，可以测试环境攻击在影响粘液清除方面的作用，并使用它们来建立清除的计算模型，这将有可能实现毒素的计算机内测试。这个有针对性的两年项目的目标是启动细胞培养和数学建模方面的技术进步，这将有助于实现有效、高效和生理上准确的毒性分析的愿景。基于细胞培养的清除试验与电子计算模型相结合的可用性将减少对动物模型的依赖，并更准确地预测毒素对人类健康的后果。

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (06) Enabling Technologies and specific Challenge Topic 06-ES-102*: 3-D or virtual models to reduce use of animals in research: Creation of miniature multi-cellular organs for high throughput screening for chemical toxicity testing. Development of novel micro-scale systems of multiple cell types that replicate the macro-scale structure and function of major organ systems in response to environmental stressors linked with development of computational models of organ system function can accelerate testing of the multitude of chemicals in our environment for toxicity. Research which furthers the generation of 3-D biological models will provide new assays for rapid screening of toxicity in organs such as the lung and liver. Cell types, such as human stem cells, used in these systems would reduce the use of animals and improve our assessment of chemical hazards in the environment. Contact: Dr. David Balshaw, balshaw@niehs.nih.gov, (919) 541-2448 Title: Computational and Cell Culture Models of Mucus Clearance. Summary: The lung inhales over 1 million infectious and toxic agents each day. The defense of the lung begins with the layer of mucus that lines the epithelial cell layer. When these assaults land at the mucus/air inter- face, the race between infection and clearance begins. The particulates, pathogens and allergens are in a diffusive race for the cell membrane before they can be cleared by the entraining flow of mucus. To provide effective clearance, the body must accomplish several goals. First it must provide the barrier layer of mucus with a height and viscoelasticity that impose a significant transit time for the infectious agents. Second, propulsion mechanisms through cilia or airflow must move the mucus with a speed that clears the agents before they can diffuse to the cell membrane. Third, the viscoelasticity of the mucus must be sufficient to prevent flooding of the airway, yet be fluid enough to allow for transport by cilia and airflow. The goal of this project is to generate computational and cell culture based models for mucus clearance. Since failure of clearance can be under- stood as the first step to a cascade of lung failure scenarios, the establishment of effective model systems for toxicity testing is critical. The use of a cell culture based model allows the inclusion of complex biochemical and immunological responses. The use of ciliated cultures that have been shown to generate and maintain appropriate mucus layers, and to generate a coordinated array of cilia for propulsion, is the starting point for our model system. We will generate the first ciliated cell culture systems that place the cells into a geometry that replicates essential features of the lung: directional, linear flow with converging cross section that can be challenged for vertical transport against gravity. This model system will have the potential to act as a sensitive as- say for environmental effectors that compromise mucus height regulation, mucus rheology, cilia density and co- ordination. We will further develop the assay within a microfluidic system that has twelve isolated assays operating in parallel. This will bring this sophisticated cell based assay to medium-throughput screening. Beyond biophysical models, we require a computational model so that we may predict the consequences of environmental assaults and design effective therapies. We will develop theoretical models that in- corporate the propulsion mechanisms of cilia and of airflow. The latter is operative during effective clearance maneuvers such as cough, and it is understood that even in the absence of cilia propulsion, cough can maintain sterile airways. However, there is currently no predictive model of the role of mucus rheology and layer thickness, cilia effectiveness and airflow in producing sufficient clearance to maintain healthy lung function. By combining a team of researchers from Applied Mathematics, Physics, Biochemistry and Biophysics, and the UNC Cystic Fibrosis Center, the goal of this project is to develop cell-based biophysical assays that can test environmental assaults for their role in compromising mucus clearance, and use them to establish a computational model for clearance that will have the potential for creating in-silico testing of toxins. The goal of this targeted two year project is to jump-start technical advances, in cell cultures and mathematical modeling, that will contribute to the vision of effective, efficient and physiologically accurate toxicity assays. The availability of a cell culture-based clearance assay coupled with an in- silico computational model will reduce the reliance on animal models and more accurately predict the consequences of toxins on human health.