State Key Laboratory of Emerging Infectious Diseases, The University of Hong Kong, Hong Kong.
Department of Microbiology, The University of Hong Kong, Pokfulam, Hong Kong.
Proc Natl Acad Sci U S A. 2018 Jun 26;115(26):6822-6827. doi: 10.1073/pnas.1806308115. Epub 2018 Jun 11.
Novel reassortant avian influenza H7N9 virus and pandemic 2009 H1N1 (H1N1pdm) virus cause human infections, while avian H7N2 and swine H1N1 virus mainly infect birds and pigs, respectively. There is no robust in vitro model for assessing the infectivity of emerging viruses in humans. Based on a recently established method, we generated long-term expanding 3D human airway organoids which accommodate four types of airway epithelial cells: ciliated, goblet, club, and basal cells. We report differentiation conditions which increase ciliated cell numbers to a nearly physiological level with synchronously beating cilia readily discernible in every organoid. In addition, the differentiation conditions induce elevated levels of serine proteases, which are essential for productive infection of human influenza viruses and low-pathogenic avian influenza viruses. We also established improved 2D monolayer culture conditions for the differentiated airway organoids. To demonstrate the ability of differentiated airway organoids to identify human-infective virus, 3D and 2D differentiated airway organoids are applied to evaluate two pairs of viruses with known distinct infectivity in humans, H7N9/Ah versus H7N2 and H1N1pdm versus an H1N1 strain isolated from swine (H1N1sw). The human-infective H7N9/Ah virus replicated more robustly than the poorly human-infective H7N2 virus; the highly human-infective H1N1pdm virus replicated to a higher titer than the counterpart H1N1sw. Collectively, we developed differentiated human airway organoids which can morphologically and functionally simulate human airway epithelium. These differentiated airway organoids can be applied for rapid assessment of the infectivity of emerging respiratory viruses to human.
新型重配禽流感 H7N9 病毒和大流行 2009 年 H1N1（H1N1pdm）病毒引起人类感染，而禽流感 H7N2 和猪流感 H1N1 病毒主要分别感染鸟类和猪。目前还没有稳健的体外模型来评估新兴病毒在人类中的感染性。基于最近建立的方法，我们生成了长期扩增的 3D 人呼吸道类器官，其中容纳了四种类型的呼吸道上皮细胞：纤毛细胞、杯状细胞、分泌细胞和基底细胞。我们报告了能使纤毛细胞数量增加到几乎生理水平的分化条件，每个类器官中都能看到同步跳动的纤毛。此外，分化条件还诱导了丝氨酸蛋白酶水平的升高，这对于人流感病毒和低致病性禽流感病毒的有效感染至关重要。我们还建立了改进的 2D 单层分化气道类器官培养条件。为了证明分化的气道类器官能够识别具有人类感染力的病毒，我们将 3D 和 2D 分化的气道类器官应用于评估两对具有已知不同人类感染性的病毒，即 H7N9/Ah 与 H7N2 以及 H1N1pdm 与从猪中分离出的 H1N1 株（H1N1sw）。具有人类感染力的 H7N9/Ah 病毒比具有较差人类感染力的 H7N2 病毒复制更为活跃；具有高度人类感染力的 H1N1pdm 病毒的复制滴度高于相应的 H1N1sw。总之，我们开发了分化的人呼吸道类器官，其在形态和功能上能够模拟人呼吸道上皮。这些分化的气道类器官可用于快速评估新兴呼吸道病毒对人类的感染性。
