Increased peripheral blood neutrophil activation phenotypes and NETosis in critically ill COVID-19 patients: a case series and review of the literature

Masso-Silva, Jorge A; Moshensky, Alexander; Lam, Michael T Y; Odish, Mazen; Patel, Arjun; Xu, Le; Hansen, Emily; Trescott, Samantha; Nguyen, Celina; Kim, Roy; Perofsky, Katherine; Perera, Samantha; Ma, Lauren; Pham, Josephine; Rolfsen, Mark; Olay, Jarod; Shin, John; Dan, Jennifer M; Abbott, Robert; Ramirez, Sydney; Alexander, Thomas H; Lin, Grace Y; Fuentes, Ana Lucia; Advani, Ira; Gunge, Deepti; Pretorius, Victor; Malhotra, Atul; Sun, Xin; Duran, Jason; Hepokoski, Mark; Crotty, Shane; Coufal, Nicole G; Meier, Angela; Alexander, Laura E Crotty

BACKGROUND: Increased inflammation has been well defined in COVID-19, while definitive pathways driving severe forms of this disease remain uncertain. Neutrophils are known to contribute to immunopathology in infections, inflammatory diseases and acute respiratory distress syndrome (ARDS), a primary cause of morbidity and mortality in COVID-19. Changes in neutrophil function in COVID-19 may give insight into disease pathogenesis and identify therapeutic targets. METHODS: Blood was obtained serially from critically ill COVID-19 patients for eleven days. Neutrophil extracellular trap formation (NETosis), oxidative burst, phagocytosis and cytokine levels were assessed. Lung tissue was obtained immediately post-mortem for immunostaining. Pubmed searches for neutrophils, lung and COVID-19 yielded ten peer-reviewed research articles in English. RESULTS: Elevations in neutrophil-associated cytokines IL-8 and IL-6, and general inflammatory cytokines IP-10, GM-CSF, IL-1b, IL-10 and TNF, were identified both at first measurement and across hospitalization (p<0.0001). COVID neutrophils had exaggerated oxidative burst (p<0.0001), NETosis (p<0.0001) and phagocytosis (p<0.0001) relative to controls. Increased NETosis correlated with leukocytosis and neutrophilia, and neutrophils and NETs were identified within airways and alveoli in lung parenchyma of 40% of SARS-CoV-2 infected lungs available for examination (2 out of 5). While elevations in IL-8 and ANC correlated with disease severity, plasma IL-8 levels alone correlated with death. CONCLUSIONS: Literature to date demonstrates compelling evidence of increased neutrophils in the circulation and lungs of COVID-19 patients. importantly, neutrophil quantity and activation correlates with severity of disease. Similarly, our data shows that circulating neutrophils in COVID-19 exhibit an activated phenotype with enhanced NETosis and oxidative burst.