ACE2 Expression in Organotypic Human Airway Epithelial Cultures and Airway Biopsies.

Qianyu Chen,Xumei Gao,Nadeene Clarke,Sinéad M Williams,Matthew J Gartner,Michelle L Baker,Yuxiu C Xia,Elizabeth A Pharo,Shawn Todd,Alastair G Stewart,Sarath Ranganathan,Kanta Subbarao,Shenna Langenbach,Meina Li

doi:10.3389/fphar.2022.813087

Abstract

Coronavirus disease 2019 (COVID-19) caused by infection with the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an acute respiratory disease with systemic complications. Therapeutic strategies for COVID-19, including repurposing (partially) developed drugs are urgently needed, regardless of the increasingly successful vaccination outcomes. We characterized two-dimensional (2D) and three-dimensional models (3D) to establish a physiologically relevant airway epithelial model with potential for investigating SARS-CoV-2 therapeutics. Human airway basal epithelial cells maintained in submerged 2D culture were used at low passage to retain the capacity to differentiate into ciliated, club, and goblet cells in both air-liquid interface culture (ALI) and airway organoid cultures, which were then analyzed for cell phenotype makers. Airway biopsies from non-asthmatic and asthmatic donors enabled comparative evaluation of the level and distribution of immunoreactive angiotensin-converting enzyme 2 (ACE2). ACE2 and transmembrane serine proteinase 2 (TMPRSS2) mRNA were expressed in ALI and airway organoids at levels similar to those of native (i.e., non-cultured) human bronchial epithelial cells, whereas furin expression was more faithfully represented in ALI. ACE2 was mainly localized to ciliated and basal epithelial cells in human airway biopsies, ALI, and airway organoids. Cystic fibrosis appeared to have no influence on ACE2 gene expression. Neither asthma nor smoking status had consistent marked influence on the expression or distribution of ACE2 in airway biopsies. SARS-CoV-2 infection of ALI cultures did not increase the levels of selected cytokines. Organotypic, and particularly ALI airway cultures are useful and practical tools for investigation of SARS-CoV-2 infection and evaluating the clinical potential of therapeutics for COVID-19.

Highlights

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)
Bronchial brushings of ciliated cells, goblet cells, club cells, and basal cells were fixed prior to ex vivo culture and cell types identified by cell-specific markers: acetylated tubulin, MUC5AC, CC10, p63, and KRT5 respectively (Supplementary Figure S1B)
air-liquid interface culture (ALI) organotypic cultures proved the most promising of airway cellular models for studies of SARS-CoV-2 infection. Both ALI and airway organoid organotypic culture re-expressed angiotensin-converting enzyme 2 (ACE2) and TMPRSS2, which were suppressed in primary epithelial cell cultures under submerged conditions which lack ciliated, secretory or goblet cell phenotypes

Summary

Introduction

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Severe cases of COVID-19 can lead to hypoxia, respiratory failure, and death. Molnupiravir and paxlovid have shown significant reduction in risk of hospital admission or death (Mahase, 2021a; b). Both agents have been authorized by FDA as emergency use for the treatment of mild-to-moderate COVID-19. Control of COVID-19 is complicated by the emergence of variant viruses that are more transmissible (e.g., the delta variant, the omicron variant) or against which current vaccines show reduced effectiveness (beta variant). An inevitable fraction of the vaccinated population who fail to mount an adequate immune response and the potential development of new virus strains resistant to vaccines further emphasize the necessity to develop additional treatment approaches for COVID-19, including repurposed drugs and novel agents

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Conclusion