Use of differentiated primary human airway epithelial models to study SARS-CoV-2 infection biology and pathogenesis

Abstract

The coronavirus disease 2019 (COVID-19) global pandemic, which so far has resulted in about 1 million deaths, is caused by severe acute respiratory coronavirus 2 (SARS-CoV-2). At present, infection dynamics and pathogenesis of the disease are incompletely understood. Here, we used well-differentiated primary human tracheal and bronchial epithelial cells (PTEC/PBEC) cultured at the air-liquid interface (ALI) to study the infection kinetics and subsequent epithelial response. ALI-PTEC and PBEC were differentiated for 3, 4 or 5 weeks, and next infected with SARS-CoV-2 to assess the impact of differentiation status on replication dynamics. We observed a gradual increase in viral load with prolonged differentiation time, and therefore focused on 5-weeks differentiated cultures. Next, we assessed the epithelial response to infection, identified the epithelial cell types that are predominantly infected and studied expression of SARS-CoV-2 entry-related host factors by using various methods (PCR, viral plaque assay and immunofluorescence [IF]). We found that ALI-PTEC cultures contained more goblet cells and less ciliated cells and showed higher expression of angiotensin-converting enzyme 2 (ACE2) compared to ALI-PBEC. Despite these differences, both cultures were efficiently infected by SARS-CoV-2. IF analysis showed that SARS-CoV-2 targets both ciliated and goblet cells. Expression of SARS-CoV-2 entry-related factors (ACE2 and TMPRSS2), IL-8 and IL-6 as well as type I and III interferons were increased after infection. These findings provide a foundation to further study virus-airway epithelial cell interactions in relation to host susceptibility to COVID-19. This study was supported by a grant from ZonMw and Proefdiervrij (# 114025007).