ACE2 and Proinflammatory Signaling by S1 Protein of SARS-Cov-2 in Human Endothelial Cells

ABSTRACT

Introduction COVID-19 is primarily a respiratory disease associated with cardiovascular risk. SARS-CoV-2, the virus causing COVID-19, uses ACE2, an important enzyme in the cardiovascular system that regulates the conversion of Ang II (deleterious/pro-hypertensive) to Ang 1-7 (protective/anti-hypertensive), as a receptor for host cell entry and infection. Considering the relationship between the viral S1-protein and the host's ACE2, it is unclear whether this interaction is merely a mechanism of infection or whether it also contributes to cardiovascular damage associated with COVID-19. We hypothesisedthat SARS-Cov-2-ACE2 interaction induces activation of vascular cell inflammatory responses that are influenced by ACE2 dependent and/or independent enzymatic Ang-(1-7) production. Methods Human microvascular endothelial cells (MEC) were used and stimulated with SARS-CoV-2 recombinant S1 protein (rS1p) (0.66 ?g/mL) at 10/30 min (acute) and 5/24h (chronic). Activation of pro-inflammatory signaling pathways (immunoblotting, real-time PCR), microparticle (MP) generation (NanoSight), and cytokine production (ELISA) were assessed. In some experiments, cells were pre-incubated with an ACE2 activator (DIZE ? 190 nM) and inhibitor (MLN-4760 ? 440 pM). Results rS1P increased NF?B activation (Control ?=0.99±0.06 vs. 1.38±0.19 AU;p<0.05) and MP formation (C=1.01±0.17 vs. 2.06±0.21, x109/mL;p<0.05), a marker of endothelial cell damage. mRA expression of IL-1? (C=1.07±0.13 vs. 50.04±4.63 2