Circulating epithelial cell as viral infection and tissue origin marker in patients with severe COVID-19

Abstract

Liquid biopsy (LB) is a minimally invasive procedure that detects biomarkers in body fluids for real-time monitoring of patients. This study developed a new LB approach to analyze Circulating Epithelial Cells (CECs) in Intensive Care Unit (ICU) patients with severe COVID-19 and High-Exposure Negative Population to COVID-19 (HENPC) as the control group. The CECs were characterized by multispectral imaging flow cytometry, and an anti-SARS-CoV-2 Spike S1 protein (ProtS) antibody was used to detect infection. The results showed that CECs were present in most ICU patients (p = 0.0412), and their median number was significantly higher (p = 0.0004) than in controls. CEC clusters were only identified in patients, and high positive ProtS expression was observed in CECs from ICU patients compared to negative controls. In conclusion, LB could be a minimally invasive tool for detecting tissue damage caused by infectious agents and could provide real-time biological information about disease status and evolution. However, further validation in a larger population of patients is needed.

Keywords: Circulating epithelial cell; Intensive care unit; Liquid biopsy; Protein S; SARS-CoV2.

Graphical abstract

Fig. 1

Cell line characterization and patient phenotyping with the Amnis ImageStream MKII Imaging System. A-upper panel: CD8 positive expression (red) on the Jurkat cell line. A-middle panel: CD14 positive (purple) expression on healthy donors. A-lower panel: Cytokeratin (CK) expression (orange) in hAELVi cells. Nuclear staining (7-AAD) (blue) determines cell integrity, a requirement to define an epithelial cell. (B) Images showing in green the presence of SARS-CoV-2 Spike S1 protein (ProtS) in patient cells. The nucleus was identified using a 7AAD fluorescent intercalator able to bind to DNA (blue). Identification of the other cell types was performed with anti-cytokeratin (CK) in orange for epithelial cells, anti-CD14 in purple for macrophages, and anti-CD8 in red for lymphocytes. ProtS was detected in Intensive Care Unit (ICU) patients in cells expressing only CK. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 2

Circulating epithelial cells (CECs) were identified based on cytokeratin (CK) expression. (A) Scatter plot of percentage of CK positiveness in High-Exposure Negative Population to COVID-19 (HENPC) controls and Intensive Care Unit (ICU) patients, with medians and interquartile ranges (p-value∗∗∗∗ ​= ​0.0004). (B) Amnis ImageStream MKII Imaging picture with cytokeratin (CK) in orange and nuclear staining showing a cluster. (C) CEC positive for RAGE expression, and D) CEC positive for ASGR1 expression. Both, specific markers of lung and liver tissue, respectively. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)