key: cord-0850898-1xc0ny1u
authors: Valle Martins, Ana Luiza; da Silva, Filipe Alex; Bolais-Ramos, Lucas; de Oliveira, Gisele Capanema; Ribeiro, Renata Cunha; Pereira, Danilo Augusto Alves; Annoni, Filippo; Diniz, Mirella Monique Lana; Silva, Thuanny Granato Fonseca; Zivianni, Bruna; Cardoso, Alexandre Carvalho; Martins, Juliana Carvalho; Motta-Santos, Daisy; Campagnole-Santos, Maria José; Taccone, Fabio Silvio; Verano-Braga, Thiago; Santos, Robson Augusto Souza
title: Increased circulating levels of angiotensin-(1–7) in severely ill COVID-19 patients
date: 2021-08-02
journal: ERJ Open Res
DOI: 10.1183/23120541.00114-2021
sha: 738c42204e0b5713208ece9b6a54710f10de7a02
doc_id: 850898
cord_uid: 1xc0ny1u

This letter reports an unexpected increase of the ACE2 product angiotensin-(1–7) and a parallel decrease of its substrate angiotensin II, suggesting a dysregulation of the renin–angiotensin system towards angiotensin-(1–7) formation in #COVID19 patients https://bit.ly/3xFXuTU

ERJ OPEN RESEARCH RESEARCH LETTER A.L. VALLE MARTINS ET AL. acid (concentration factor 20×) and analysed by LC-MS/MS (Xevo TQ-S, Waters) using the multiple reaction monitoring mode (figure 1c). The calibration curve was obtained using a stock solution containing synthetic peptides (Bachem) of all RAS peptides used in this study. The applied calibration curve model ( y=ax+b) proved accurate over the concentration range from 10 to 1000 pg·mL −1 (r=0.997). The limit of quantitation, and inter-and intravariability of this method have been previously reported [7] . Data are presented as mean±SEM and the parametric t-test was used for the statistical analyses. Although we did not measure the tissular levels of RAS peptides, our findings contrast with the initial hypothesis that the interaction of SARS-CoV-2 with ACE2 would result in higher Ang II and lower Ang-(1-7) levels compared to non-COVID-19 subjects [4] . Recent studies using the equilibrium method to measure the ACE2 activity [8, 9] are in line with the results presented here, as they reported higher Ang-(1-7) and lower Ang II plasma levels in severe COVID-19. For general clinical studies including nonsevere COVID-19 patients, it seems that all circulating RAS peptides are reduced due to decreased activity of renin [10] . The role of ACE2 in the observed results is questionable as previous studies suggested that one of the main routes to produce Ang-(1-7) in the circulation is by ACE2-independent pathways [11, 12] . The observed significant decrease of Ang II and increase of Ang-(1-7) arterial levels in severe COVID-19 patients (figure 1d) is probably due to a direct dysregulation of RAS pathways in COVID-19 rather than a direct consequence of ACEi usage (figure 1e). The Ang-(1-7)/Ang II ratio, which is an estimation of Ang II→Ang-(1-7) conversion, was three-fold higher in COVID-19 patients (2.79±0.682 versus 0.878±0.201; p=0.0141), which may suggest an increased ACE2 or other Ang-(1-7)-forming activity in COVID-19. Increased soluble ACE2 in severe COVID-19 patients have been recently reported [9, 13, 14] . REINDL-SCHWAIGHOFER et al. [9] reported that ACE2 level increased over the course of the disease, reaching its maximum peak after ∼10 days of hospitalisation. Increased ACE2 correlated with increased Ang-(1-7) and decreased Ang II levels [9] , suggesting its important role in controlling the circulating RAS peptides in severe COVID-19.

In contrast to a previous report [3] , we observed a significant reduction of Ang II concentration in COVID-19 patients, which may add more data against the reliability of ELISA to measure Ang II in human plasma [5] . The Ang II/Ang I ratio was not significantly altered in the COVID-19 patients (0.205±0.0322 versus 0.292±0.0517; p=0.1518), which may suggest that ACE activity is not altered in COVID-19. Unfortunately, we were unable to perform direct measurements of enzymatic activities in this study due to the denaturating conditions that we used to collect the samples, but our estimation of increased ACE2 activity in severe COVID-19 blood samples is in line with previous reports [9, 14] . The remaining question to be answered, though, is whether the observed increased Ang-(1-7) and decreased Ang II levels are a direct effect of SARS-CoV-2 or a consequence of inflammation due to the infection that triggers Ang-(1-7) synthesis via Ang II metabolisation.

Limitations of our study includes the small cohort included (n=19 for each group) and the lack of direct activity measurements of the RAS-related enzymes. Nonetheless, this is the first report of the direct measurement of RAS peptides in the arterial circulation of severe COVID-19 patients. Although future studies are obviously necessary to better understand the effects of this disease on RAS pathways, our data provide new insights for the interpretation and planning of future therapies to modulate the RAS in the context of COVID-19.

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Ang II (angiotensin II) conversion to angiotensin-(1-7) in the circulation is POP (prolyloligopeptidase)-dependent and ACE2 (angiotensin-converting enzyme 2)-independent

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Conflict of interest: A.L. Valle Martins reports a patent pending (BR1020210009950). F.A. da Silva has nothing to disclose. L. Bolais-Ramos has nothing to disclose. G.C. de Oliveira has nothing to disclose. R.C. Ribeiro has nothing to disclose. D.A.A. Pereira has nothing to disclose. F. Annoni has nothing to disclose. M.M.L. Diniz has nothing to disclose. T.G.F. Silva has nothing to disclose. B. Ziviani has nothing to disclose. A.C. Cardoso has nothing to disclose. J.C. Martins has nothing to disclose. D. Motta-Santos has nothing to disclose. M.J. Campagnole-Santos has nothing to disclose. F.S. Taccone has nothing to disclose. T. Verano-Braga has nothing to disclose. R.A.S. Santos reports patents planned.