Neutrophils can promote clotting via FXI and impact clot structure via neutrophil extracellular traps in a distinctive manner in vitro

Y Shi,J S Gauer,S D Connell,R A S Ariëns,S R Baker,H Philippou

doi:10.1038/s41598-021-81268-7

Abstract

Neutrophils and neutrophil extracellular traps (NETs) have been shown to be involved in coagulation. However, the interactions between neutrophils or NETs and fibrin(ogen) in clots, and the mechanisms behind these interactions are not yet fully understood. In this in vitro study, the role of neutrophils or NETs on clot structure, formation and dissolution was studied with a combination of confocal microscopy, turbidity and permeation experiments. Factor (F)XII, FXI and FVII-deficient plasmas were used to investigate which factors may be involved in the procoagulant effects. We found both neutrophils and NETs promote clotting in plasma without the addition of other coagulation triggers, but not in purified fibrinogen, indicating that other factors mediate the interaction. The procoagulant effects of neutrophils and NETs were also observed in FXII- and FVII-deficient plasma. In FXI-deficient plasma, only the procoagulant effects of NETs were observed, but not of neutrophils. NETs increased the density of clots, particularly in the vicinity of the NETs, while neutrophils-induced clots were less stable and more porous. In conclusion, NETs accelerate clotting and contribute to the formation of a denser, more lysis resistant clot architecture. Neutrophils, or their released mediators, may induce clotting in a different manner to NETs, mediated by FXI.

Highlights

Neutrophils and neutrophil extracellular traps (NETs) have been shown to be involved in coagulation
A key function via which neutrophils contribute to innate immune responses and to thrombosis is the generation of neutrophil extracellular traps (NETs), which are released by activated neutrophils, and contain DNA and granular proteins[21,22]
Previous studies have shown that neutrophils and NETs are involved in both arterial and venous thrombosis[33,34,35,36,37], questions remain whether neutrophils contribute to coagulation independently or via NETs, whether both neutrophils and NETs promote clotting via the intrinsic pathway and how procoagulant factors interact with neutrophils or NETs

Summary

Introduction

Neutrophils and neutrophil extracellular traps (NETs) have been shown to be involved in coagulation. The interactions between neutrophils or NETs and fibrin(ogen) in clots, and the mechanisms behind these interactions are not yet fully understood In this in vitro study, the role of neutrophils or NETs on clot structure, formation and dissolution was studied with a combination of confocal microscopy, turbidity and permeation experiments. Factor (F)XII, FXI and FVII-deficient plasmas were used to investigate which factors may be involved in the procoagulant effects We found both neutrophils and NETs promote clotting in plasma without the addition of other coagulation triggers, but not in purified fibrinogen, indicating that other factors mediate the interaction. The role of neutrophils or NETs on clot formation and clot dissolution was analysed by turbidity and lysis assays, clot structure was determined by confocal microscopy, and clot stability and pore size were investigated using permeation studies We found that both partially activated neutrophils and NETs promote clotting in plasma independently of additional triggers. The procoagulant effects of neutrophils are mediated via FXI, to a lesser extent by FXII, but not by FVII

Methods

Results

Conclusion