Bioengineered Multi-Cell Type Organoids For Airways Disease Modeling

项目摘要 呼吸道疾病是全世界的主要死因之一。引起的呼吸系统疾病 吸烟、感染和遗传因素加在一起造成每年约950万人死亡。 呼吸道感染是世界范围内第三大最常见的死亡原因，也是世界上主要的死亡原因。 发展中国家遗传风险因素是肺部疾病患病率的重要因素。比如说， 囊性纤维化（CF）被列为最广泛的缩短寿命的遗传性疾病之一2 - 4， 目前有7万人患有CF。由于发病率高，呼吸道疾病也是 大多数研究的医疗条件。呼吸系统疾病的研究由于缺乏合适的 在体内和体外模型中研究呼吸道上皮、感染和 疾病不幸的是，肺部疾病的动物模型在气道发育方面与人类显著不同 和疾病病理学，所以经常导致不准确和有明显缺陷的模型，虽然一些进展 已经在使用人类细胞培养系统进行疾病建模和药物发现，目前在体外 模型不能再现复杂的空间形态，并允许生物学相关的细胞-细胞和 细胞-基质相互作用拟议工作的总体前提是：（a）体内动物模型通常不同 在疾病病理学中显著地来自人类，并且具有显著的成本限制;和（B）目前的体外 呼吸道疾病和细菌发病机理的模型不能概括复杂的组织成分 和人体气道上皮的3D结构。这个合作R01提案的动机是 迫切需要解决目前的差距和知识，并克服目前体外2D细胞的局限性， 用于气道疾病建模、治疗开发和临床前测试的培养模型。我们的整体 假设体内气道微环境重现将提供更有效的体外 用于气道疾病建模和治疗评估的替代物。为了验证这一假设，我们将首先生成 生物工程化多细胞3D气道类器官，由肺细胞外基质（ECM）衍生的支架支持 具有可调的生物力学特性，以促进健康气道的多细胞组织和功能 上皮（Aim 1）。接下来，我们将评估含有CF气道上皮的3D气道类器官是否可以 模型疾病病理学（目的2）。最后，我们将使用这种新的气道疾病模型来研究 铜绿假单胞菌的发病机制（目的3）。如果获奖，这项合作R 01将使我们的多学科和 多机构团队开发和评估3D气道类器官作为更有效的体外替代物， 气道疾病建模和治疗评估。在这项工作的基础上可能取得的进展包括使用 用于测试个性化疾病治疗的患者特异性气道类器官模型， 收集临床数据，以验证类器官的使用，以预测治疗结果并指导治疗。

PROJECT SUMMARY Respiratory diseases are among the leading causes of death worldwide. Respiratory disease caused by smoking, infections, and genetic factors together account for approximately 9.5 million deaths per year. Respiratory infection is the 3rd most common cause of death worldwide, and the leading cause of death in developing countries. Genetic risk factors are significant contributors to lung disease prevalence. For example, cystic fibrosis (CF) is ranked as one of the most widespread life-shortening genetic diseases2-4 with more than 70,000 people currently living with CF. Due to the high incidence; respiratory diseases are also among the most studied medical conditions. The study of respiratory diseases is significantly limited by a lack of suitable in vivo and in vitro models to investigate interactions between the respiratory epithelium, infection, and disease. Unfortunately animal models of lung disease differ significantly from humans in airway development and disease pathology, so often result in inaccurate and significantly flawed models, While some progress has been made in using human cell culture systems for disease modeling and drug discovery, current in vitro models are unable to reproduce the complex spatial morphology and allow biologically relevant cell-cell and cell-matrix interactions. The overall premise for the proposed work is that (a) in vivo animal models often differ significantly from humans in disease pathology and have significant cost limitations; and (b) current in vitro models of respiratory disease and bacterial pathogenesis do not recapitulate the complex tissue components and 3D architecture of the human airway epithelium. This collaborative R01 proposal is motivated by the critical need to address the current gaps and knowledge and overcome the limitations of current in vitro 2D cell culture models for airway disease modeling, therapy development, and pre-clinical testing. Our overall hypothesis is that recapitulation of the in vivo airway microenvironment will provide a more effective in vitro surrogate for airway disease modeling and therapy evaluation. To test this hypothesis, we will first generate bioengineered multicellular 3D airway organoids, supported by a lung extracellular matrix (ECM)-derived biogel with tunable biomechanical properties, to promote multicellular organization and function of healthy airway epithelium (Aim 1). Next we will evaluate whether 3D airway organoids containing CF airway epithelium can model disease pathology (Aim 2). Finally we will use this novel airway disease model to study the pathogenesis of P. aeruginosa (Aim 3). If awarded, this collaborative R01 will allow our multi-disciplinary and multi-institutional team to develop and evaluate 3D airway organoids as a more effective in vitro surrogate for airway disease modeling and therapy evaluation. Likely developments building on this work could include use of patient-specific airway organoid models for testing of personalized disease treatments, combined with collection of clinical data to validate the use of organoids to predict treatment outcomes and guide treatment.