Defective NET clearance contributes to sustained FXII activation in COVID-19-associated pulmonary thrombo-inflammation

Abstract

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.

Identification of the factor XII contact activation site enables sensitive coagulation diagnostics

“…Western blotting was performed using primary goat polyclonal anti-FXII antibody (1:1000) and HRP-coupled anti-goat antibody (1:5000) 51 . 20 μL of the incubated material was diluted in buffer (1:6) and analysed using the chromogenic substrate S2302 (1 mM) at an absorbance wavelength of 405 nm in a Bio-Kinetics Reader (SpectraMax Plus, Molecular Devices) at 37 °C 69 . Samples activated with PKa were pre-incubated with aprotinin (100 KIU/mL).…”

The contact system in liver injury

“…Moreover, clinical trials are currently performed with FXIIa-blocking antibodies CSL312/garadacimab (CSL Behring). Targeting FXIIa might be beneficial for diseases with aberrant FXIIa activity, e.g., the kallikrein–kinin system in HAE patients and thrombosis-related complications in SARS-CoV-2 infections [ 97 ]. Especially, the absence of bleeding tendency by FXIIa blockade may support its use as a safe antithrombotic treatment in liver disease because conventional direct oral anticoagulants that prevent frequently occurring PVT in early-stage disease will become inapplicable in patients with late-stage chronic liver disease [ 98 ].…”

Sex-specific differences in plasma levels of FXII, HK, and FXIIa-C1-esterase inhibitor complexes in community-acquired pneumonia

“…4) (36,42). This assessment becomes particularly true in the context of COVID-19 pathology, in which immense injury of endothelial cells and sustained activation of FXII lead to excessive microvascular thrombosis (43,44). In addition, the sex-specific effects in health and disease have to be recognized when studying the interplay between coagulation and immune responses in the context of lung pathologies.…”