Distinct temporal characteristics of circulating alveolar epithelial and endothelial injury markers in ARDS with COVID-19

© The Author(s) 2021. Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http:// creat iveco mmons. org/ licen ses/ by/4. 0/. The Creative Commons Public Domain Dedication waiver (http:// creat iveco mmons. org/ publi cdoma in/ zero/1. 0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. In the most severe cases, coronavirus disease (COVID19) leads to acute respiratory distress syndrome (ARDS) that is characterised by alveolar epithelial and endothelial injuries [1]. There are several ARDS biomarkers, which reflect alveolar tissue injuries [2]. The level of circulating soluble form of receptor for advanced glycation end-products (sRAGE) is correlated with type-1 alveolar epithelial injuries [3]. The elevation of angiopoietin-2 (ANG-2) indicates endothelial injury in patients with ARDS [4]. The increase in the circulating surfactant protein concentration indicates alveolar barrier disruption in ARDS cases [5]. Recently, Spadaro et al. reported that COVID-19 ARDS is characterised by increase in the circulating endothelial injury markers [6]. However, the detailed temporal characteristics of these markers remain unclear. In this preliminary study, we investigated the levels of circulating sRAGE, ANG-2, and surfactant protein D (SP-D) in serum samples of patients with COVID-19 with or without ARDS. Patients who were diagnosed with COVID-19 by realtime polymerase chain reaction and admitted to Yokohama City University Hospital from January to August 2020 were included in this retrospective observational study (Ethics Reference Number: B200700100). Serum concentrations of sRAGE, ANG-2, and SP-D were measured using enzyme-linked immunosorbent assay kits (human RAGE: DY1145; human ANG-2: DY623; human SP-D: DY1920; R&D systems, Minneapolis, MN, USA). We compared the concentrations of these markers in patients with and without ARDS on hospital day 1 or 2. Moreover, we analysed temporal changes in these markers during the first 8 hospital days in those with ARDS. ARDS was diagnosed according to the Berlin Definition. The data of those with and without ARDS were compared with the Mann–Whitney U test. Temporal changes in the markers were analysed using the Friedman and post-hoc Dunn’s tests. The peak day for each biomarker was observed using the Kruskal–Wallis and post-hoc Dunn’s tests. All statistical analyses were performed using Prism 9.0 software (Graphpad Software, San Diego CA, USA). The level of significance was set at P < 0.05. Eleven and ten patients with and without ARDS, respectively, all with COVID-19, were included. Their characteristics are presented in Table 1. ARDS diagnosis was made on hospital day 1 or 2. The initial serum levels of sRAGE and SP-D were significantly higher in the ARDS than in the non-ARDS group; however, no significant difference was observed in the ANG-2 levels (Table 1). Analysis of temporal changes in these markers in 10 patients with ARDS, after excluding one patient with missing data, revealed that the serum sRAGE level peaked just after admission, and gradually decreased Open Access

In the most severe cases, coronavirus disease (COVID-19) leads to acute respiratory distress syndrome (ARDS) that is characterised by alveolar epithelial and endothelial injuries [1]. There are several ARDS biomarkers, which reflect alveolar tissue injuries [2]. The level of circulating soluble form of receptor for advanced glycation end-products (sRAGE) is correlated with type-1 alveolar epithelial injuries [3]. The elevation of angiopoietin-2 (ANG-2) indicates endothelial injury in patients with ARDS [4]. The increase in the circulating surfactant protein concentration indicates alveolar barrier disruption in ARDS cases [5]. Recently, Spadaro et al. reported that COVID-19 ARDS is characterised by increase in the circulating endothelial injury markers [6]. However, the detailed temporal characteristics of these markers remain unclear. In this preliminary study, we investigated the levels of circulating sRAGE, ANG-2, and surfactant protein D (SP-D) in serum samples of patients with COVID-19 with or without ARDS.
Patients who were diagnosed with COVID-19 by realtime polymerase chain reaction and admitted to Yokohama City University Hospital from January to August 2020 were included in this retrospective observational study (Ethics Reference Number: B200700100). Serum concentrations of sRAGE, ANG-2, and SP-D were measured using enzyme-linked immunosorbent assay kits (human RAGE: DY1145; human ANG-2: DY623; human SP-D: DY1920; R&D systems, Minneapolis, MN, USA). We compared the concentrations of these markers in patients with and without ARDS on hospital day 1 or 2. Moreover, we analysed temporal changes in these markers during the first 8 hospital days in those with ARDS. ARDS was diagnosed according to the Berlin Definition.
The data of those with and without ARDS were compared with the Mann-Whitney U test. Temporal changes in the markers were analysed using the Friedman and post-hoc Dunn's tests. The peak day for each biomarker was observed using the Kruskal-Wallis and post-hoc Dunn's tests. All statistical analyses were performed using Prism 9.0 software (Graphpad Software, San Diego CA, USA). The level of significance was set at P < 0.05.
Eleven and ten patients with and without ARDS, respectively, all with COVID-19, were included. Their characteristics are presented in Table 1. ARDS diagnosis was made on hospital day 1 or 2. The initial serum levels of sRAGE and SP-D were significantly higher in the ARDS than in the non-ARDS group; however, no significant difference was observed in the ANG-2 levels ( Table 1).
Analysis of temporal changes in these markers in 10 patients with ARDS, after excluding one patient with missing data, revealed that the serum sRAGE level peaked just after admission, and gradually decreased with hospital days (Fig. 1). Conversely, serum ANG-2 and SP-D levels did not significantly decrease during the first 8 hospital days and the peak timings of these markers were observed during a later disease stage (Fig. 1).
We showed that alveolar epithelial injury occurring at the very early disease stage, indicated by the increased sRAGE level, is a hallmark of COVID-19 ARDS. Conversely, the ANG-2 and SP-D levels peaked at later time points, suggesting that the endothelial injury and alveolar barrier disruption continued to exacerbate for several days after admission.
A study limitation was that the precise mechanism of sRAGE or ANG-2 release from the alveolar epithelial or endothelial cells remains unknown. However, the difference in the peak timing of these markers suggested distinct mechanisms for injury to each cell type. Additionally, it is possible that the initial alveolar epithelial injury might be a trigger of the subsequent exacerbation. Further investigations analysing temporal associations between these markers and inflammatory mediators could help identify the mechanisms underlying alveolar tissue injury. Moreover, the trajectory analysis of these markers linking clinical outcomes could help understand the detailed COVID-19 pathogenesis.