A team of scientists from Australia and Brazil recently investigated the involvement of endothelial cells in the pathogenesis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. The study reveals that endothelial cells are less susceptible to SARS-CoV-2 infection unless there is high viral load in the blood or epithelial basolateral infection. However, these cells can significantly contribute to SARS-CoV-2 pathogenesis by inducing proinflammatory responses. The study is currently available on the bioRxiv* preprint server.
SARS-CoV-2, the causative pathogen of coronavirus disease 2019 (COVID-19), has been found to cause severe pulmonary and cardiovascular complications, particularly in susceptible individuals. Being a respiratory virus, SARS-CoV-2 primarily attacks pulmonary epithelial cells before encountering pulmonary endothelial cells. In the context of COVID-19-related cardiovascular pathologies, some autopsy studies have indicated the presence of viral RNA in the vascular network of many organs, whereas some studies have failed to show any direct endothelial infection by SARS-CoV-2.
Because endothelial cells are known to express angiotensin-converting enzyme 2 (ACE2), which is the host cell receptor for SARS-CoV-2 entry, it is theoretically possible to induce endothelial SARS-CoV-2 infection. A growing-pool of evidence suggests that SARS-CoV-2 infection-induced disruption of the pulmonary epithelial barrier may result in direct endothelial infection via the basolateral side of the epithelial layer, which is located adjacent to the pulmonary endothelial layer.
In the current study, the scientists aim to investigate whether COVID-19-related vascular pathologies are caused by direct endothelial infection or inflammation-induced activation of the endothelium.
To investigate the SARS-CoV-2-induced alterations in the endothelial cells in vivo, the scientists collected lung tissue samples from 10 deceased COVID-19 patients and stained for the spike mRNA of SARS-CoV-2. Moreover, they conducted a series of in vitro studies using primary human endothelial cells and an experimental model of the pulmonary epithelial-endothelial cell barrier to evaluate SARS-CoV-2 pathogenesis.
According to the in vivo findings, no spike mRNA was detected in the lung tissues. This indicates that SARS-CoV-2 does not directly infect pulmonary endothelial cells. Moreover, the findings of the in vitro experimentations revealed that primary human endothelial cells expressed mRNAs of both ACE2 and TMPRSS2 (host cell protease required for spike priming) and that the levels of expressions were comparable to that observed in human epithelial cell line (Calu-3). However, at the protein level, significantly reduced expressions of ACE2 and TMPRSS2 were observed in endothelial cells compared to that in epithelial cells. These observations indicate that compared to epithelial cells, endothelial cells are less susceptible to SARS-CoV-2 infection because of lower expression levels of ACE2 and TMPRSS2.
To further investigate the possibility of endothelial infection, the scientists experimentally infected primary endothelial cells with SARS-CoV-2 either apically or basolaterally. An apical infection occurs when the virus is present in the bloodstream, and a basolateral infection occurs when the virus is present in the basolateral surface of the adjacent epithelial layer. However, in both the infection conditions, they failed to detect any sign of viral replication within these cells. Moreover, they failed to detect SARS-CoV-2 nuclear protein in these cells. Importantly, no considerable morphological alteration was observed in endothelial cells after SARS-CoV-2 infection.
By increasing the amount of SARS-CoV-2 for experimental infection, they noticed a significantly increased expression of viral nuclear protein in endothelial cells. However, viral replication remained undetected. These observations indicate that although SARS-CoV-2 can enter endothelial cells when present in high concentrations, it cannot replicate to induce productive infection.
To investigate inflammatory responses of SARS-CoV-2-infected endothelial cells, they measured the expression of intercellular adhesion molecule 1, which is known to be induced in endothelial cells in response to inflammation. In addition, they assessed whether infected endothelial cells secrete proinflammatory cytokines. By conducting immunofluorescence staining and biochemical assays, they observed increased expression of intercellular adhesion molecule 1 and robust secretion of interleukin 6 (IL-6) in endothelial cells infected with SARS-CoV-2 either apically or basolaterally. These observations indicate that although causing an abortive infection, SARS-CoV-2 can induce inflammatory responses in endothelial cells.
By infecting a more realistic co-culture model of the lung epithelial-endothelial barrier with SARS-CoV-2, they observed that unlike epithelial cells, which clearly displayed the signs of infection, endothelial cells failed to express viral nuclear protein or RNA upon infection. With further analysis, they observed that endothelial cells responded to the infection in the nearby epithelial cells by inducing the secretion of proinflammatory mediators.
The study reveals that endothelial cells are susceptible to SARS-CoV-2 infection only when the virus is present in the nearby basolateral epithelial surface or when present in the circulation in a significantly high amount. Even if SARS-CoV-2 enters endothelial cells, the infection is only associated with inflammatory responses and not with viral amplification.
bioRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.