Development of 3D Alveolar Tissue Models, CRISPR-editing and Microbiota-immune Response Assay Platforms for Deciphering Human Lung Immunity

技术开发项目 技术开发项目（Tech Dev）的目标是开发方法、工具和测定， 满足杰克逊实验室人类免疫学合作中心（JAX CCHI）的需求， 提高科学界解决人类肺免疫、人类免疫和人类免疫系统等问题的能力， 免疫-微生物群相互作用和免疫细胞的基本机制。肺免疫相关的主要问题 功能仍然没有答案-例如免疫和肺上皮细胞之间的细胞与细胞的相互作用， 形状对外来因子的反应，或者气道或肺室内微生物群的存在如何 影响病毒感染和其他肺部疾病的发病机制。研究的一个重大技术障碍 人类免疫-肺动力学是人类肺的绝对复杂性-它不断过滤空气 通过肺上皮和居民之间的动态相互作用， 免疫细胞如巨噬细胞或树突细胞。这种复杂性在动物模型中很难模拟 系统或使用死亡的人类肺组织。为了克服这些挑战，Tech Dev将专注于三个方面 创新人体组织平台：1）肺部和上呼吸道的三维（3D）生物打印模型 研究暴露于病毒或代谢性疾病期间功能性肺免疫相互作用组的环境 2）基于CRISPR/Cas9的工具，用于对原代人免疫细胞亚群、造血细胞亚群和免疫细胞亚群进行基因工程改造; 用于探测细胞功能的干细胞和/或肺上皮祖细胞;和3）用于 筛选肺部驻留微生物群并确定它们对人类肺部免疫应答的影响。中的每 这些平台解决了应用程序中未满足的特定需求，并将使我们能够分别研究 在动态和生理相关微环境中的人肺免疫，以询问特定的 预测对病毒感染作出反应的细胞类型和分子途径，并评估细菌感染的影响。 从人气道分离的代谢物对抗病毒反应的影响。通过这些努力，JAX CCHI将 装备，以解决以前无法解决的问题，有关肺免疫动力学，朝着一个更 对肺免疫功能的机制性理解。我们的具体目标是： 目标1。使用3D生物打印开发人类肺组织-免疫相互作用的体外模型。 目的2：优化基于CRISPR的遗传工具，用于工程化原代人类免疫和肺上皮细胞 细胞 目的3：建立一个功能性免疫分析平台，以确定人类免疫调节的景观 肺和气道微生物群。

PROJECT SUMMARY TECHNOLOGY DEVELOPMENT PROJECT The goal of the Technology Development project (Tech Dev) is to develop approaches, tools and assays that address the needs of The Jackson Laboratory Cooperative Center on Human Immunology (JAX CCHI) and that advance the capabilities of the scientific community to tackle questions regarding human lung immunity, human immune-microbiota interactions and basic mechanisms of immune cells. Major questions related to lung immune function remain unanswered—such as the cell-to-cell interactions between immune and lung epithelial cells that shape responses to foreign agents, or how the presence of microbiota in the airways or within lung compartments influences the pathogenesis of viral infections and other lung diseases. A significant technical barrier to studying human immune-lung dynamics is the sheer complexity of the human lung—which constantly filters airborne particles, infectious organisms and air through dynamic interactions between the lung epithelium and resident immune cells such as macrophages or dendritic cells. This complexity cannot be easily modeled in animal systems or using deceased human lung tissue. To surmount these challenges, Tech Dev will focus on three innovative human tissue platforms: 1) three-dimensional (3D) bioprinted models of the lung and upper respiratory environment for investigating the functional lung-immune interactome during exposure to viral or metabolic agents; 2) CRISPR/Cas9-based tools to genetically engineer primary human immune cell subsets, hematopoietic stem cells and/or lung epithelial progenitors to probe cell function; and 3) a functional in vitro platform for screening lung-resident microbiota and determining their impact on human lung immune responses. Each of these platforms addresses a specific unmet need in the application and will enable us, respectively, to study the human lung immunity within a dynamic and physiologically relevant microenvironment, to interrogate specific cell types and molecular pathways predicted to respond to viral infections, and to assess the impact of bacterial metabolites isolated from human airways on antiviral responses. Through these efforts, the JAX CCHI will be equipped to address previously inaccessible questions related to lung-immune dynamics, towards a more mechanistic understanding of lung immune function. Our Specific Aims are: Aim 1. Develop in vitro models of human lung tissue-immune interactions using 3D bioprinting. Aim 2: Optimize CRISPR-based genetic tools for use in engineering primary human immune and lung epithelial cells. Aim 3: Develop a functional immune assay platform to determine the immunomodulatory landscape of human lung and airway microbiota.