Embedded Printing of Human Respiratory Model with Air-Liquid Interface for COVID-19 Research

项目摘要 本研究的总体目标是制造体外三维（3D）人体呼吸道 具有气液界面（ALI）的模型，可作为COVID-19相关生物医学的平台 调查事务所最近爆发的COVID-19疫情是由于感染了严重急性呼吸道感染， 综合征冠状病毒2（SARS-CoV-2），导致超过1亿例确诊病例和约230万例 全球死亡人数每天都在增加。对于COVID-19相关研究，仍然存在紧迫的问题， 我们需要一个完全异质的3D细胞呼吸模型，这可以增强我们对呼吸系统的理解。 SARS-CoV-2诱导肺损伤并促进新治疗方法的开发，作为对动物的补充 模型和临床患者。因此，我们建议开发这样的人体呼吸模型，其中1） 表达血管紧张素转换酶2（ACE 2）蛋白和跨膜蛋白酶丝氨酸2型 （TMPRSS 2）允许病毒进入研究SARS-CoV-2感染过程，以及2）具有扩散性 气道和主流体通道之间的渗透性，以使质量运输穿过ALI， 筛选临床证明有效的COVID-19药物。我们假设体外人类呼吸系统 模型可以通过嵌入式打印和进一步开发屈服应力中的灌注结构来实现 含有原代人基质细胞、内皮细胞和成纤维细胞的基质浴。 因此，提出了以下两个具体目标： 目的1：在基于基质细胞的3D体外人呼吸模型中的嵌入式打印和灌注 交叉屈服应力基体浴。 目的2：通过检测ACE 2受体和TMPRSS 2来表征人呼吸模型 气道上皮细胞中的蛋白酶和测定ALI的扩散渗透性。 首次提出了一种基于明胶微凝胶和明胶溶液的交叉屈服应力细胞 复合基质浴将被创新用于其中可灌注组织构造的嵌入式打印， 人体呼吸模型制作。该模型开发研究将产生体外3D人体 利用新的交叉屈服应力细胞基质建立的ALI和细胞基质呼吸模型 浴缸的嵌入式打印，没有其他模型与这种生理复杂性存在。这样的人类 呼吸模型将为研究COVID-19大流行相关感染提供一个通用的体外平台 加工和筛选相关药物。

Project Summary The overarching goal of this research is to fabricate an in vitro three-dimensional (3D) human respiratory model with an air-liquid interface (ALI), which can serve as a platform for COVID-19 related biomedical investigations. A recent COVID-19 outbreak, which is due to the infection of a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), results in more than 100 million confirmed cases and about 2.3 million deaths with numbers increasing everyday globally. For COVID-19 related research, there remains an urgent need for a 3D fully heterogeneous, cellular respiratory model, which can enhance our understanding of how SARS-CoV-2 induces lung injury and facilitate the development of new treatments as complementary to animal models and clinical patients. We therefore propose to develop such a human respiratory model, which 1) expresses angiotensin-converting enzyme 2 (ACE2) protein and transmembrane protease serine type 2 (TMPRSS2) that allow viral entry to study the SARS-CoV-2 infection process, and 2) has the diffusional permeability between the airway and primary fluid channel to enable the mass transport across ALI, permitting the screening of clinically proven drugs for COVID-19 use. We hypothesize that an in vitro human respiratory model can be realized by embedded printing and further developing a perfusable construct in a yield-stress matrix bath containing primary human stromal, endothelial, and fibroblast cells. Accordingly, two specific aims are proposed as follows: Aim 1: Embedded printing and perfusion of a 3D in vitro human respiratory model in a stromal cell-based cross-linkable yield-stress matrix bath. Aim 2: Characterization of the human respiratory model via detection of ACE2 receptor and TMPRSS2 protease in the airway epithelium and determination of the diffusional permeability of ALI. For the first time, a gelatin microgels and gelatin solution-based cross-linkable yield-stress cellular composite matrix bath will be innovated for embedded printing of perfusable tissue constructs in it, enabling human respiratory model fabrication. This model development study will result in an in vitro 3D human respiratory model with ALI and cellular stroma by utilizing the novel cross-linkable yield-stress cellular matrix bath for embedded printing, and no other models with this physiological complexity exist. Such a human respiratory model will provide a versatile in vitro platform for studying the COVID-19 pandemic-related infection process and screening related drugs.