Repressed Ang 1-7 in COVID-19 Is Inversely Associated with Inflammation and Coagulation

Abstract

The coronavirus SARS-CoV-2 infects host cells by binding to the angiotensin-converting enzyme 2 (ACE2) receptor, which belongs to an anti-inflammatory, anti-thrombotic counter-regulatory arm of the renin-angiotensin system (RAS). ACE2 dysfunction and RAS dysregulation has been explored as a driving force in acute respiratory distress syndrome (ARDS), but data from COVID-19 patients has been inconsistent and inconclusive. We sought to identify disruptions of the classical (ACE)/angiotensin (Ang) II/Ang II type-1 receptor (AT₁R) and the counter-regulatory ACE2/Ang 1-7/Mas Receptor (MasR) pathways in patients with COVID-19 and correlate these with severity of infection and markers of inflammation and coagulation. Ang II and Ang 1-7 levels in plasma were measured by enzyme-linked immunosorbent assay (ELISA) for 230 patients, 166 of whom were SARS-CoV-2+. Ang 1-7 was repressed in COVID-19 patients compared to that in SARS-CoV-2 negative outpatient controls. Since the control cohort was less sick than the SARS-CoV-2+ group, this association between decreased Ang 1-7 and COVID-19 cannot be attributed to COVID-19 specifically as opposed to critical illness more generally. Multivariable logistic regression analyses demonstrated that every 10-pg/mL increase in plasma Ang 1-7 was associated with a 3% reduction in the odds of hospitalization (adjusted odds ratio [AOR] 0.97, confidence interval [CI] 0.95 to 0.99) and a 3% reduction in odds of requiring oxygen supplementation (AOR 0.97, CI 0.95 to 0.99) and/or ventilation (AOR 0.97, CI 0.94 to 0.99). Ang 1-7 was also inversely associated with pro-inflammatory cytokines and d-dimer in this patient cohort, suggesting that reduced activity in this protective counter-regulatory arm of the RAS contributes to the hyper-immune response and diffuse coagulation activation documented in COVID-19. IMPORTANCE Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes a unique disease, COVID-19, which ranges in severity from asymptomatic to causing severe respiratory failure and death. Viral transmission throughout the world continues at a high rate despite the development and widespread use of effective vaccines. For those patients who contract COVID-19 and become severely ill, few therapeutic options have been shown to provide benefits and mortality rates are high. Additionally, the pathophysiology underlying COVID-19 disease presentation, progression, and severity is incompletely understood. The significance of our research is in confirming the role of renin-angiotensin system dysfunction in COVID-19 pathogenesis in a large cohort of patients with diverse disease severity and outcomes. Additionally, to our knowledge, this is the first study to pair angiotensin peptide levels with inflammatory and thrombotic markers. These data support the role of ongoing clinical trials examining renin-angiotensin system-targeted therapeutics for the treatment of COVID-19.

Keywords: Ang 1–7; COVID-19; coagulation; inflammation; renin-angiotensin system.

FIG 1

Schematic representation of the systemic renin-angiotensin system. Renin converts angiotensinogen into angiotensin I (Ang I). Angiotensin-converting enzyme (ACE) converts Ang I into angiotensin II (Ang II), which binds to Ang II type 1 receptor (AT₁R), through which it exerts its harmful inflammatory effects. Angiotensin-converting enzyme 2 (ACE2) converts the majority of Ang II to Ang 1–7, which activates the Mas receptor (MasR) signaling pathway with protective downstream effects on the microcirculatory environment. Attachment of SARS-CoV-2 spike protein to ACE2 induces cleavage and release of the soluble form of ACE2 by ADAM-17, virus-receptor complex internalization and receptor downregulation. Reduced ACE2 expression is hypothesized to cause an imbalance between the classical ACE/Ang I/AT₁R and the protective ACE2/Ang 1–7/MasR axes that is central to the pathophysiology of coronavirus disease 2019 (COVID-19).

FIG 2

Reduced ACE2 activity evidenced by repressed Ang 1–7 and increased Ang II:Ang 1–7 ratios. (A and B) Boxplots show the distribution of Ang 1–7 and Ang II levels (pg/mL) stratified by COVID-19 status and adverse outcomes, including hospitalization, need for oxygen supplementation, ventilation, and death. Above each outcome is the unadjusted P value from nonparametric Wilcoxon Mann-Whitney U tests. The numbers of unique patient samples quantified are as follows: Angiotensin II (n = 202), Angiotensin 1–7 (n = 229). (C) The log-transformed ratio of Ang II to Ang 1–7 is displayed. t test P values are included above each boxplot from analyzing each outcome category separately. *, P < 0.05; **, P < 0.005; ***, P < 0.0005.

FIG 3

Inverse relationship of Ang II and Ang 1–7 to mean arterial blood pressures (MAP) (A and B) Linear regression between Ang II and Ang 1–7 levels and MAP averaged from all available readings taken on the day of sample collection. (C and D) Linear regression between Ang II and Ang 1–7 and potassium levels collected from the day of sample collection. The numbers of measurements available on the day of sample collection are as follows: MAP (n = 156 mm Hg), Potassium (mEq/L) (n = 98).

FIG 4

Correlation between angiotensin peptides, d-dimer, and pro-inflammatory cytokines in COVID-19. Correlation matrix depicts the Spearman’s correlation coefficient between angiotensin 1–7, angiotensin II, d-dimer, and cytokine levels on a colorimetric scale from negative correlation in red to positive correlation in blue. The cytokines displayed were selected based on significant associations with Angiotensin 1–7 levels using Kruskal-Wallis tests and analyzing each cytokine separately. The numbers of unique patient samples quantified are as follows: all cytokines (n = 164), angiotensin II (n = 202), angiotensin 1–7 (n = 229), d-dimer retrospectively pulled from clinical records (n = 54), MAP (n = 156).