Self-organized stem cell-derived human lung buds with proximo-distal patterning and novel targets of SARS-CoV-2

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the global COVID-19 pandemic and the lack of therapeutics hinders pandemic control. Although lung disease is the primary clinical outcome in COVID-19 patients, how SARS-CoV-2 induces tissue pathology in the lung remains elusive. Here we describe a high-throughput-based platform to generate tens of thousands of self-organizing, nearly identical, and genetically matched human lung buds derived from human pluripotent stem cells (hPSCs) cultured on confined geometries on micropattern chips. Strikingly, in vitro-derived human lung buds resemble fetal human lung tissue and display in vivo-like proximo-distal coordination of alveolar and airway tissue differentiation whose 3D epithelial self-organization is directed by the levels of KGF. Single-cell transcriptomics unveiled the cell identities and ontogeny of airway and alveolar tissue and the specification of WNThi cycling alveolar stem cells from alveolar progenitors. These synthetic human lung buds are susceptible to SARS-CoV-2 infection and can be used to track cell type-dependent susceptibilities to infection, intercellular transmission and cytopathology in airway and alveolar tissue in individual synthetic lung buds. We detected an increased susceptibility to infection in alveolar cells and identified cycling alveolar stem cells as targets of SARS-CoV-2. We used this platform to test neutralizing antibodies isolated from convalescent plasma that efficiently blocked SARS-CoV-2 infection and intercellular transmission. Our platform offers unlimited, rapid and scalable access to disease-relevant lung tissue that recapitulate human lung development and can be used to track SARS-CoV-2 infection and identify pre-clinical candidate therapeutics for COVID-19. Overall design: Examination of cell type composition and heterogeneity in synthetic human lungs.

严重急性呼吸综合征冠状病毒2型（SARS-CoV-2）是引发全球新型冠状病毒肺炎（Corona Virus Disease 2019，COVID-19）大流行的病原体，治疗手段的匮乏阻碍了疫情防控工作。尽管肺部疾病是COVID-19患者的主要临床转归，但SARS-CoV-2诱导肺部组织病理损伤的具体机制仍未阐明。本研究报道了一种基于高通量技术的培养平台，可在微图案芯片的受限几何培养环境中，由人类多能干细胞（human pluripotent stem cells，hPSCs）批量培养生成数以万计的自组织、高度均一且遗传背景一致的人肺芽。令人惊喜的是，体外诱导获得的人肺芽与胎儿人肺组织形态相似，且呈现出体内样的肺泡与气道组织分化的近-远轴协调模式，其三维上皮自组织过程受角质细胞生长因子（Keratinocyte Growth Factor，KGF）水平调控。单细胞转录组测序揭示了气道与肺泡组织的细胞身份及发育起源，并阐明了肺泡祖细胞向WNT高表达（WNThi）的循环肺泡干细胞的特化过程。这些人工合成的人肺芽易受SARS-CoV-2感染，可用于追踪单个合成肺芽中气道与肺泡组织内的细胞类型依赖性感染易感性、细胞间传播及细胞病理变化。本研究检测到肺泡细胞的感染易感性升高，并确定循环肺泡干细胞是SARS-CoV-2的感染靶标。我们利用该平台测试了从康复者血浆中分离的中和抗体，其可有效阻断SARS-CoV-2的感染与细胞间传播。本平台可实现无限、快速且规模化的疾病相关肺组织获取，该组织能够复现人类肺发育过程，可用于追踪SARS-CoV-2感染并筛选COVID-19的临床前候选治疗药物。整体实验设计：对人工合成人肺组织的细胞类型组成及异质性进行检测分析。