Hypoxia alters the expression of ACE2 and TMPRSS2 SARS-CoV-2 cell entry mediators in hCMEC/D3 brain endothelial cells

Abstract

The mechanisms by which the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) induces neurological complications remain to be elucidated. We aimed to identify possible effects of hypoxia on the expression of SARS-CoV-2 cell entry mediators, angiotensin-converting enzyme 2 (ACE2) receptor and transmembrane protease serine 2 (TMPRSS2) protein, in human brain endothelial cells, in vitro . hCMEC/D3 cells were exposed to different oxygen tensions: 20% (Control group), 8% or 2% O 2 (Hypoxia groups). Cells were harvested 6-, 24- and 48 h following hypoxic challenge for assessment of mRNA and protein, using qPCR and Western Blot. The response of the brain endothelial cells to hypoxia was replicated using modular incubator chambers. We observed an acute increase (6 h, p < 0.05), followed by a longer-term decrease (48 h, p < 0.05) in ACE2 mRNA and protein expression, accompanied by reduced expression of TMPRSS2 protein levels (48 h, p < 0.05) under the more severe hypoxic condition (2% O 2 ). No changes in levels of von Willebrand Factor (vWF – an endothelial cell damage marker) or interleukin 6 (IL-6 – a pro-inflammatory cytokine) mRNA were observed. We conclude that hypoxia regulates brain endothelial cell ACE2 and TMPRSS2 expression in vitro , which may indicate human brain endothelial susceptibility to SARS-CoV-2 infection and subsequent brain sequelae.

SARS-CoV-2 infection of human brain microvascular endothelial cells leads to inflammatory activation through NF-κB non-canonical pathway and mitochondrial remodeling

“…cells(Imperio et al, 2021). These observations are in accordance with our present data, that ACE2 is decreased while HIF-1α is increased at 24 hpi.…”

Long-Term Sequelae of COVID-19 in Experimental Mice

“…While viral infection may directly impact CNS cells that affect these factors, possibly by stimulating hypoxic/ischemic and hemorrhagic lesions [ 48 ], these findings may not be specific to direct viral infection on the brain since these effects could be secondary to systemic inflammation and coagulopathy caused by viral infection. In support of the possible direct effect of the virus on the brain, a few studies have shown (1) the presence of SARS-CoV-2 mRNA in cerebrospinal fluid of patients with acute COVID-19 [ 49 , 50 ], (2) the presence of ACE2 receptors in the blood–brain barrier (BBB) [ 51 , 52 ], and (3) altered BBB integrity and reduced blood flow, as well as the presence of SARS-CoV-2 in cortical neurons of patients who died of COVID-19 and immune cell infiltrates [ 48 , 53 – 55 ]. Since these findings have only been reported in acute stages following SARS-CoV-2 infection and given that these changes may occur either directly or indirectly, it is still unclear how viral infection impacts the brain over a longer period.…”

SARS-CoV-2 infection of human brain microvascular endothelial cells leads to inflammatory activation through NF-κB non-canonical pathway and mitochondrial remodeling

“…HIF-1α is activated and translocated to the nucleus upon hypoxic conditions (Ke & Costa, 2006) and it has been shown that COVID-19 patients present massive hypoxia due to vasoconstriction and coagulopathy (Afsar et al, 2020). Interestingly, ACE2 expression is decreased in pulmonary smooth muscle cells upon HIF-1α accumulation (Zhang et al, 2020), whereas hypoxia leads to a biphasic modulation of both ACE2 and TMPRSS2 expression on brain microvascular endothelial cells (hCMEC/D3), with an initial increase at 6 h and a decrease at 48 h of hypoxic stimulus cells (Imperio et al, 2021). These observations are in accordance with our present data, that ACE2 is decreased while HIF-1α is increased at 24 hpi.…”