Proteomics identifies a type I IFN, prothrombotic hyperinflammatory circulating COVID-19 neutrophil signature distinct from non-COVID-19 ARDS

Summary 

This study reports on the proteomics and metabolomics analysis of peripheral blood neutrophils from patients with COVID-19 ARDS (CA; n=3), non-COVID-19 ARDS (NA; n=3-7), moderate COVID-19 (MC; n=3), and healthy controls (HC; n=4-7). For some analyses normal density neutrophils (NDN) and low density neutrophils (LDN; isolated from the PBMC layer) were analysed separately. LDN consisted of both mature (CD16+CD10+) and immature (CD16-CD10-) neutrophils. Due to the small samples sizes most conclusions will need further validation, but the data suggest that neutrophils in COVID-19 ARDS may have increased IFN I signalling, degranulation, and platelet binding compared to healthy controls. 

Research Highlights  

1. Comparing the neutrophil proteome of COVID-19 ARDS patients to healthy controls highlights an increase in the type I interferon signalling pathway and in markers of platelet degranulation. 

2. Mature LDN from moderate or ARDS COVID-19 patients show increased CD41 staining, suggesting increased binding to platelets, compared to NDN from healthy controls. Immature LDN from COVID-19 patients do not show increased CD41 staining. 

3. NDN from ARDS COVID-19 patients have slightly decreased content of granule proteins compared to healthy controls. 

4. NDN from ARDS COVID-19 patients have increased glucose content compared to healthy controls, the cause is unclear. 

5. In vitro stimulation of healthy neutrophils with a TLR7/8 agonist under hypoxic conditions leads to an activated phenotype regarding surface receptors. 

Impact for COVID-19 research:  

No direct impact because the conclusions need further validation. 

Methodologies: 

• Study Type: case study, in vitro 

• Key Techniques: flow cytometry for activation markers, proteomics, metabolomics, extracellular flux analysis 

Limitations: 

• Small group sizes; n=3 for COVID-19 ARDS and n=3 for moderate COVID-19. 

• p-hacking obvious: the grouping & subsetting of the samples is not consistent throughout the manuscript, and comparisons are often illogical (e.g. why not compare NA directly to CA in Fig. 3?). 

• Most statements in the text are based on comparisons that don’t show statistically significant differences. 

• Patient characteristics are only provided for ARDS patients not for moderate patients or HC. Unclear whether non-COVID-19 ARDS patients also had viral infection.