Background: Coagulopathy and inflammation are hallmarks of Coronavirus disease 2019 and are associated with increased mortality. Clinical and experimental data have revealed a role for neutrophil extracellular traps (NETs) in COVID-19 disease. The mechanisms that drive thrombo-inflammation in COVID-19 are poorly understood. Methods: We performed proteomic analysis and immunostaining of postmortem lung tissues from COVID-19 patients and patients with other lung pathologies. We further compared coagulation factor XII (FXII) and DNase activities in plasma samples from COVID-19 patients and healthy control donors and determined NET-induced FXII activation using a chromogenic substrate assay. Findings: FXII expression and activity were increased in the lung parenchyma, within the pulmonary vasculature and in fibrin-rich alveolar spaces of postmortem lung tissues from COVID-19 patients. In agreement with this, plasmaaac acafajf€ oeFXII activation (FXIIa) was increased in samples from COVID-19 patients. Furthermore, FXIIa colocalized with NETs in COVID-19 lung tissue indicating that NETs accumulation leads to FXII contact activation in COVID-19. We further showed that an accumulation of NETs is partially due to impaired NET clearance by extracellular DNases as DNase substitution improved NET dissolution and reduced FXII activation in vitro. Interpretation: Collectively, our study supports that the NET/FXII axis contributes to the pathogenic chain of procoagulant and proinflammatory responses in COVID-19. Targeting both NETs and FXIIa may offer a potential novel therapeutic strategy.

Coagulopathy and inflammation are hallmarks of Coronavirus disease 2019 (COVID-19) and are associated with increased mortality. The mechanisms that drive thrombo-inflammation in COVID-19 are poorly understood. Here, we report a role of the NETs/ Factor XII (FXII) axis for initiating procoagulant and proinflammatory reactions in COVID-19.Proteomic analysis revealed enrichment of FXII in postmortem lung tissue from COVID-19 patients. Immunofluorescence analysis of COVID-19 lung tissue showed that FXII is activated in the lung parenchyma, within the pulmonary vessel walls and in fibrin-rich alveolar spaces. In particular, activated FXII (FXIIa) colocalized with NETs in COVID-19 lung tissue, indicating that NETs accumulation leads to FXII contact activation in COVID-19. We further showed that an accumulation of NETs is partially caused by impaired NETs clearance via extracellular DNases. In contrast, addition of DNase I improved NETs clearance and reduced FXII activation in vitro. We propose that targeting both, FXIIa and the FXII activator NETs, is therapeutically effective in mitigating thrombo-inflammation in COVID-19.

Contact activation refers to the process of surface-induced activation of factor XII (FXII), which initiates blood coagulation and is captured by the activated partial thromboplastin time (aPTT) assay. Here, we show the mechanism and diagnostic implications of FXII contact activation. Screening of recombinant FXII mutants identified a continuous stretch of residues Gln317–Ser339 that was essential for FXII surface binding and activation, thrombin generation and coagulation. Peptides spanning these 23 residues competed with surface-induced FXII activation. Although FXII mutants lacking residues Gln317–Ser339 were susceptible to activation by plasmin and plasma kallikrein, they were ineffective in supporting arterial and venous thrombus formation in mice. Antibodies raised against the Gln317–Ser339 region induced FXII activation and triggered controllable contact activation in solution leading to thrombin generation by the intrinsic pathway of coagulation. The antibody-activated aPTT allows for standardization of particulate aPTT reagents and for sensitive monitoring of coagulation factors VIII, IX, XI.