Endothelial cells elicit a pro-inflammatory response to SARS-CoV-2 without productive viral infection

Objectives Thrombotic and microvascular complications are frequently seen in deceased COVID-19 patients. However, whether this is caused by direct viral infection of the endothelium or inflammation-induced endothelial activation remains highly contentious. Methods Here, we use patient autopsy samples, primary human endothelial cells and an in vitro model of the pulmonary epithelial-endothelial cell barrier to show that primary human endothelial cells express very low levels the SARS-CoV-2 receptor ACE2 and the protease TMPRSS2. Results Accordingly, endothelial cells can only be infected when SARS-CoV-2 is present at very high concentrations. However, this is not a productive infection (i.e. no infectious virus is produced) and viral entry induces an inflammatory response. We also show that SARS-CoV-2 does not infect endothelial cells in 3D vessels under flow conditions. We further demonstrate that in a co-culture model endothelial cells are not infected with SARS-CoV-2. They do however sense and respond to infection in the adjacent epithelial cells, increasing ICAM-1 expression and releasing pro-inflammatory cytokines. Conclusions Taken together, these data suggest that in vivo , endothelial cells are unlikely to be infected with SARS-CoV-2 and that infection is only likely to occur if the adjacent pulmonary epithelium is denuded (basolateral infection) or a high viral load is present in the blood (apical infection). In such a scenario, whilst SARS-CoV-2 infection of the endothelium can occur, it does not contribute to viral amplification. However, endothelial cells are still likely to play a key role in SARS-CoV-2 pathogenesis by sensing adjacent infection and mounting a pro-inflammatory response to SARS-CoV-2. ### Competing Interest Statement KS is a co-inventor on patent applications for NLRP3 inhibitors which have been licensed to Inflazome Ltd, a company headquartered in Dublin, Ireland. Inflazome is developing drugs that target the NLRP3 inflammasome to address unmet clinical needs in inflammatory disease. KS served on the Scientific Advisory Board of Inflazome in 2016 - 2017, and serves as a consultant to Quench Bio, USA and Novartis, Switzerland.