Defective NET clearance contributes to sustained FXII activation in COVID-19-associated pulmonary thrombo-inflammation
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Abstract
Background
Coagulopathy and inflammation are hallmarks of Coronavirus disease 2019 (COVID-19) 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 Factor XIII (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, plasma FXII 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 NETs/FXII axis contributes to the pathogenic chain of procoagulant and proinflammatory responses in COVID-19. Targeting both, NETs and FXIIa, could provide a strategy to mitigate COVID-19-induced thrombo-inflammation.
Funding
This study was supported by the European Union (840189), the Werner Otto Medical Foundation Hamburg (8/95) and the German Research Foundation (FR4239/1-1, A11/SFB877, B08/SFB841 and P06/KFO306).
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SciScore for 10.1101/2020.12.29.424644: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement IRB: Acquisition of patient plasma samples: Heparinized plasma samples were obtained from SARS-CoV-2 positive patients hospitalized at the University Medical Center Hamburg, Germany during the first outbreak in April 2020 (permit number #2322, Ethics Committee of the Hamburg Medical Association, Hamburg, Germany). Randomization The experiments were not randomized. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
Antibodies Sentences Resources The following antibodies were used: anti-neutrophil elastase (5 μg/ml, ab121595, Abcam) anti-neutrophil elastase ( 5suggested: NoneSoftware and Algorithms Sentences Resources T… SciScore for 10.1101/2020.12.29.424644: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement IRB: Acquisition of patient plasma samples: Heparinized plasma samples were obtained from SARS-CoV-2 positive patients hospitalized at the University Medical Center Hamburg, Germany during the first outbreak in April 2020 (permit number #2322, Ethics Committee of the Hamburg Medical Association, Hamburg, Germany). Randomization The experiments were not randomized. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
Antibodies Sentences Resources The following antibodies were used: anti-neutrophil elastase (5 μg/ml, ab121595, Abcam) anti-neutrophil elastase ( 5suggested: NoneSoftware and Algorithms Sentences Resources The diameter of DNase digestion was measured using Image J. Image Jsuggested: (ImageJ, RRID:SCR_003070)Statistics: GraphPad Prism was used for graphic representation and statistical analysis of data. GraphPadsuggested: (GraphPad Prism, RRID:SCR_002798)The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium22 via the PRIDE23 partner repository with the dataset identifier PXD020216. ProteomeXchangesuggested: (ProteomeXchange, RRID:SCR_004055)Results from OddPub: Thank you for sharing your data.
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: We found the following clinical trial numbers in your paper:
Identifier Status Title NCT04355364 Recruiting Efficacy and Safety of aerosolizedDornase Alfa Administratio… NCT04402970 Completed Dornase Alfa for ARDS in Patients With SARS-CoV-2 NCT04445285 Recruiting Phase 2 Trial Using rhDNase to Reduce Mortality in COVID-19 … NCT04432987 Recruiting Dornase Alpha for the Treatment of COVID-19 NCT04359654 Recruiting Nebulised Dornase Alfa for Treatment of COVID-19 NCT04402944 Recruiting Pulmozyme to Improve COVID-19 ARDS Outcomes NCT04524962 Recruiting Study of Descartes-30 in Acute Respiratory Distress Syndrome NCT04541979 Recruiting Aerosoliserat DNase for Treatment of Respiratory Failure in … NCT04409925 Recruiting DISmantling COvid iNduced Neutrophil ExtraCellular Traps (DI… NCT04409509 Completed Treatment With CSL312 in Adults With Coronavirus Disease 201… Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
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