Single-cell RNA-seq of air-liquid interface bronchioalveolar cells

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes coronavirus disease 2019 (COVID-19), may result in acute respiratory distress syndrome (ARDS), multi-organ failure and death. The alveolar epithelium is a major target of the virus, but representative models to study virus host interactions in more detail are currently lacking. Here, we describe a human 2D air-liquid interface culture system which was characterized by confocal-, electron-microscopy and single cell mRNA expression analysis. In this model, alveolar cells, but also basal cells and rare neuroendocrine cells, are grown from 3D self-renewing lung bud tip organoids. These cultures were readily infected by SARS-CoV-2 with mainly surfactant protein C-positive ATII-like cells being targeted. Consequently, significant viral titers were detected and mRNA expression analysis revealed induction of type I/III interferon response program. Treatment of these cultures with a low dose of interferon lambda dramatically reduced viral replication. Hence, these cultures represent an experimental model for SARS-CoV-2 infection and can be applied for drug screens. We have setup a bronchioalveolar model for studying COVID-19. scRNAseq was performed to characterize the model on air-liquid interface bronchioalveolar cells not infected with SARS-CoV-2.

引发2019冠状病毒病（coronavirus disease 2019, COVID-19）的严重急性呼吸综合征冠状病毒2（Severe acute respiratory syndrome coronavirus 2, SARS-CoV-2）可导致急性呼吸窘迫综合征（acute respiratory distress syndrome, ARDS）、多器官功能衰竭乃至死亡。肺泡上皮是该病毒的主要侵染靶点，但目前仍缺乏可用于深入解析病毒-宿主相互作用的代表性研究模型。本研究报道了一种人类二维气液界面（air-liquid interface, ALI）培养系统，并通过共聚焦显微镜、电子显微镜及单细胞mRNA表达分析对该系统完成表征。该培养体系中的肺泡细胞、基底细胞及罕见神经内分泌细胞，均来源于三维自我更新肺芽尖类器官。SARS-CoV-2可高效感染此培养体系，且主要靶向表面活性蛋白C阳性的ATII样细胞，据此可检测到显著的病毒滴度；mRNA表达分析显示，感染过程可诱导I/III型干扰素应答程序。采用低剂量λ干扰素处理该培养体系，可显著抑制病毒复制。综上，该培养体系可作为研究SARS-CoV-2感染的实验模型，适用于药物筛选。我们构建了用于研究COVID-19的细支气管肺泡模型，并通过单细胞RNA测序（single-cell RNA sequencing, scRNAseq）对未感染SARS-CoV-2的ALI细支气管肺泡细胞进行了表征分析。