Laboratory of Biomedical Embryology, Department of Health, Animal Science and Food Safety and Center for Stem Cell Research, Università degli Studi di Milano, Milan, Italy.
Academic Unit of Reproductive and Developmental Medicine, Department of Oncology and Metabolism, University of Sheffield, Sheffield, UK.
Methods Mol Biol. 2021;2273:131-138. doi: 10.1007/978-1-0716-1246-0_8.
The current coronavirus disease-19 (COVID-19) pandemic, caused by "severe acute respiratory syndrome coronavirus 2" (SARS-CoV-2), underscores the threat posed by newly emerging viruses. The understanding of the mechanisms driving early infection events, that are crucial for the exponential spread of the disease, is mandatory and can be significantly implemented generating 3D in vitro models as experimental platforms to investigate the infection substrates and how the virus invades and ravages the tissues.We here describe a protocol for the creation of a synthetic hydrogel-based 3D culture system that mimics in vitro the complex architectures and mechanical cues distinctive of the upper airway epithelia. We then expose the in vitro generated 3D nasal and tracheal epithelia to gold nanoparticles (AuNPs) that display the typical shape and size distinctive of SARS-CoV-2 and of the majority of Coronaviridae presently known.The infection platform here described provides an efficient and highly physiological in vitro model that reproduces the host-pathogen early interactions, using virus-mimicking nanoparticles, and offers a flexible tool to study virus entry into the cell. At the same time, it reduces the risk of accidental infection/spillovers for researchers, which represents a crucial aspect when dealing with a virus that is highly contagious, virulent, and even deadly.
当前由“严重急性呼吸系统综合症冠状病毒 2 型”（SARS-CoV-2）引起的冠状病毒病-19（COVID-19）大流行突显了新出现病毒所构成的威胁。了解驱动疾病快速传播的早期感染事件的机制是强制性的，通过生成 3D 体外模型作为实验平台来研究感染底物以及病毒如何入侵和破坏组织，可以显著推进这一工作。我们在此描述了一种创建基于合成水凝胶的 3D 培养系统的方案，该系统可模拟上呼吸道上皮的复杂结构和机械线索。然后，我们将体外生成的 3D 鼻和气管上皮暴露于金纳米颗粒（AuNPs），这些纳米颗粒显示出 SARS-CoV-2 和目前已知的大多数冠状病毒科的典型形状和大小。本文描述的感染平台提供了一种高效且高度生理的体外模型，使用模拟病毒的纳米颗粒再现宿主-病原体的早期相互作用，并提供了一种灵活的工具来研究病毒进入细胞的过程。同时，它降低了研究人员意外感染/溢出的风险，这在处理一种高度传染性、高毒性甚至致命的病毒时是一个关键方面。
