OC 18.4 - The role of neutrophil extracellular traps (NETs) in lower extremity artery disease (LEAD)-specific thrombosis

Sunday, July 10, 2022

11:30 AM – 11:45 AM

Location: ExCel Center, ICC Capital Suite Room 2-4

Presentation Format: In-Person

Presenting Author(s)

Manovriti Thakur, PhD

Postdoc
Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland. 2. Department for BioMedical Research, University of Bern, Switzerland.
Olten, Solothurn, Switzerland

Co-Author(s)

NA

Nico Angliker

Scientific assistant
1. Division of Angiology, Swiss Cardiovascular Center, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland. 2. Department for BioMedical Research, University of Bern, Switzerland
bern, Bern, Switzerland
MS

Mark Siegrist

Technical assistant
1. Division of Angiology, Swiss Cardiovascular Center, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland. 2. Department for BioMedical Research, University of Bern, Switzerland
Bern, Bern, Switzerland
MK

Marijke Kuijpers

Associate professor
Maastricht University
Maastricht, Limburg, Netherlands
MS

Marc Schindewolf

Head of Clinical Investigation Unit
1. Division of Angiology, Swiss Cardiovascular Center, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland. 2. Department for BioMedical Research, University of Bern, Switzerland
Bern, Bern, Switzerland
IB

Iris Baumgartner

Clinic director, Professor
1. Division of Angiology, Swiss Cardiovascular Center, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland. 2. Department for BioMedical Research, University of Bern, Switzerland
Bern, Bern, Switzerland
YD

Yvonne Döring

Professor
1. Division of Angiology, Swiss Cardiovascular Center, Inselspital, Bern University Hospital, University of Bern, Bern, Switzerland. 2. Department for BioMedical Research, University of Bern, Switzerland. 3. Institute for Cardiovascular Prevention (IPEK), Ludwig-Maximilians-University Munich (LMU), 80336 Munich, Germany. 4. DZHK (German Centre for Cardiovascular Research), Partner Site Munich Heart Alliance, 80336 Munich, Germany.
Bern, Bern, Switzerland

Background: Lower-extremity arterial disease (LEAD) is a significant health problem affecting 230 million people worldwide. Thrombotic complications are a hallmark of LEAD pathophysiology and recent genome wide associations studies have identified the risk variant Factor V Leiden specifically in LEAD emphasizing the importance of arterial thrombosis in LEAD. Arterial thrombi also consists of neutrophil extracellular traps (NETs), however, their prevalence and role in LEAD is very limited.

Aims: Explore the prevalence and role of NETs (negatively charged DNA-scaffold) in the LEAD-specific thrombus.
Establishment of the ferric chloride induction thrombosis in the femoral artery of Apoe-/- mice.
Effect of therapeutic interference with NETs on the LEAD-specific thrombi.

Methods: We measured the prevalence of NETs in LEAD versus carotid thromboendarterectomy patient samples. In our in-vitro assays, we used flow chambers to perfuse the NETs with labelled fibrinogen to enable clot formation in absence or presence of NETs inhibitors like DNase1 or Oleylamine (neutralize negatively charged DNA scaffold). Furthermore, we performed thromboelastometry assays and functional assays using similar conditions. Apoe-/- mice (WD 21 weeks) were used for FeCl3 induced thrombosis in the femoral arteries in absence and presence of NET modifying reagents. 2-photon intravital microscopy and histology was used to evaluate the role of NETs and its inhibitors in thrombus formation. For statistical analyses we have used Mann-Whitney, Kruskal-Wallis with Dunn`s multiple comparison.

Results: We saw strongly abolished fibrin clotting in presence of NET interfering agents. Correspondingly, thromboelastometry assays also revealed extended clotting time in presence of NET interference in-vitro. FeCl3-induced thrombosis in femoral artery showed significant number of NETs in the thrombi leading to arterial occlusion.

Conclusion(s): LEAD specific thrombosis differs from other arterial thrombi with significantly higher prevalence of NETs. NETs inhibition leads to depletion of thrombus formation both in-vivo and in-vitro. Therefore, preventing NETs release could represent a potential strategy to prevent LEAD-specific thrombosis.