Glycyrrhizin through liquorice intake modulates ACE2 and HMGB1 levels-A pilot study in healthy individuals with implications for COVID-19 and ARDS

Felix Buder, Simina-Ramona Selejan, Mathias Hohl, Michael Kindermann, Christian Herr, Philipp M Lepper, Robert Bals, Bernd Salzberger, Felix Mahfoud, Michael Böhm, Felix Buder, Simina-Ramona Selejan, Mathias Hohl, Michael Kindermann, Christian Herr, Philipp M Lepper, Robert Bals, Bernd Salzberger, Felix Mahfoud, Michael Böhm

Abstract

Background: Glycyrrhizin, an active component of liquorice root extract, exhibits antiviral and immunomodulatory properties by direct inhibition of the pro-inflammatory alarmin HMGB1 (High-mobility group box 1).

Objective: The aim of this study was to explore the role of liquorice intake on the viral entry receptor ACE2 (angiotensin-converting enzyme 2) and the immunoregulatory HMGB1 in healthy individuals and to explore HMGB1 expression in coronavirus disease 2019 (COVID-19) or non-COVID-19 in ARDS (acute respiratory distress syndrome patients).

Material and methods: This study enrolled 43 individuals, including hospitalised patients with i) acute respiratory distress syndrome (ARDS) due to COVID-19 (n = 7) or other underlying causes (n = 12), ii) mild COVID-19 (n = 4) and iii) healthy volunteers (n = 20). Healthy individuals took 50 g of liquorice (containing 3% liquorice root extract) daily for 7 days, while blood samples were collected at baseline and on day 3 and 7. Changes in ACE2 and HMGB1 levels were determined by Western blot analysis and enzyme-linked immunosorbent assay, respectively. Additionally, HMGB1 levels were measured in hospitalised COVID-19 patients with mild disease or COVID-19 associated acute respiratory distress syndrome (ARDS) and compared with a non-COVID-19-ARDS group.

Results: Liquorice intake significantly reduced after 7 days both cellular membranous ACE2 expression (-51% compared to baseline levels, p = 0.008) and plasma HMGB1 levels (-17% compared to baseline levels, p<0.001) in healthy individuals. Half of the individuals had a reduction in ACE2 levels of at least 30%. HMGB1 levels in patients with mild COVID-19 and ARDS patients with and without COVID-19 were significantly higher compared with those of healthy individuals (+317%, p = 0.002), but they were not different between COVID-19 and non-COVID-19 ARDS.

Conclusions: Liquorice intake modulates ACE2 and HMGB1 levels in healthy individuals. HMGB1 is enhanced in mild COVID-19 and in ARDS with and without COVID-19, warranting evaluation of HMGB1 as a potential treatment target and glycyrrhizin, which is an active component of liquorice root extract, as a potential treatment in COVID-19 and non-COVID-19 respiratory disease.

Conflict of interest statement

RB reports grants and consulting fees from AstraZenica, grants and consulting fees from Boehringer Ingelheim, personal fees from GlaxoSmithKline, consulting fees from Grifols, grants and consulting fees from Novartis, personal fees from CSL Behring, grants from German Federal Ministry of Education and Research (BMBF) Competence Network Asthma and COPD (ASCONET), grants from Sander Stiftung, grants from Schwiete Stiftung, grants from Krebshilfe, grants from Mukoviszidose eV. FM is supported by Deutsche Gesellschaft für Kardiologie (DGK), Deutsche (SFB TRR219), and Deutsche Herzstiftung. He has received scientific support from Medronic and ReCor Medical and speaker honoraria from Astra-Zeneca, Bayer, Boehringer Ingelheim, Inari, Medtronic, Merck, and ReCor Medical. MB is supported by the Deutsche Forschungsgemeinschaft (German Research Foundation; TTR 219, project number 322900939) and reports personal fees from Abbott, Amgen, Astra Zeneca, Bayer, Boehringer Ingelheim, Cytokinetics, Medtronic, Novartis, Servier and Vifor during the conduct of the study. The grants and fundings indicated are however not related to this publication. Therefore, this does not alter our adherence to PLOS ONE policies on sharing data and materials.

Figures

Fig 1. Modulation of mACE2 and HMGB1…
Fig 1. Modulation of mACE2 and HMGB1 levels after liquorice intake in healthy individuals.
A, PBMC mACE2 levels before and 7 days after liquorice ingestion. B, Representative ACE2 Western blot (upper panel) of membraneous (mACE2) and cytosolic (cACE2) fractions from PBMCs from one healthy volunteer at baseline and after 7 days (7d-FU) of liquorice ingestion. The corresponding Ponceau Red strip showing total protein loading is presented below (lower panel). C, Plasma HMGB1 levels before as well as 3 and 7 days after liquorice ingestion. Wilcoxon test and Friedman analysis with Dunn’s post-hoc test were performed when comparing 2 groups and 3 groups respectively. Data are shown as median±IQR. Abbreviation: mACE2, membranous ACE2; HMGB1, high-mobility group box 1 protein; PBMC, peripheral blood mononuclear cell; IQR, interquartile range.
Fig 2. HMGB1 levels in healthy individuals,…
Fig 2. HMGB1 levels in healthy individuals, in COVID-19 patients with mild or severe course (ARDS) and in ARDS patients without COVID-19.
HMGB1 levels were evaluated in healthy individuals (n = 20), COVID-19 patients with a mild (n = 4) course and ARDS (n = 7) respectively, as well as ARDS patients without COVID-19 (n = 12). Data are shown as median±IQR. Kruskal-Wallis ANOVA with Dunn’s post-hoc test was performed. Abbreviation: ARDS, acute respiratory distress syndrome; COVID-19, Coronavirus disease 2019; HMGB1, high-mobility group box 1 protein; IQR, interquartile range.

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