OC 67.5 - Neutrophils critically influence thrombus growth and architecture by enhanced neutrophil-platelet interactions and their reciprocal recruitment during sepsis.

Background: Thrombosis and thromboembolic events are hallmark findings of sepsis, despite thrombocytopenia, platelet dysfunction and anti-platelet therapy. During the last decade, the interplay of platelets and immune cells was shown to be a powerful driver of thrombosis, yet the role of neutrophil-platelet crosstalk in immunothrombosis remains underinvestigated.

Aims: Deciphering the importance of neutrophil-platelet interaction for initiation of immunothrombosis in sepsis.

Methods: Seventy sepsis patients were recruited from the ICU at the University Hospital Würzburg (Ethical vote 92/19 63/20). A thorough analysis of thrombus formation, thrombus architecture, platelet and leukocyte phenotyping, and their interaction under static and flow conditions was conducted using an in vitro microfluidic chamber, multicolor flow cytometry, and confocal imaging. Results were compared to healthy controls.

Results: In contrast to the reduced platelet surface area coverage (SAC) found by perfusion of blood at arterial shear, venous shear revealed a 57% increase in platelet SAC (p < 0.0001) in sepsis whole blood (Fig.1A-D). We identified a 22x increase in mean leukocyte count (p < 0.0001) and total thrombus volume (Fig.1E-F). Neutrophils were identified as a critical mediator of immunothrombosis, as demonstrated by the reversion of elevated platelet SAC levels to healthy controls when either neutrophils were depleted or neutrophil-platelet interplay was inhibited by antibodies blocking PSGL-1 on neutrophils (Fig.2A-C). When neutrophils or platelets were cross-incubated, only sepsis neutrophils were increasingly recruited into the growing thrombus, while platelets were not differentially incorporated. The enhanced adhesion of neutrophils during sepsis led to a more efficient clearance of S.aureus from the thrombus (Fig.2D). Inhibition of platelet crosstalk or platelet depletion reverted this finding, emphasizing the importance of the neutrophil-platelet interface for infection control.

Conclusion(s): Targeting the platelet-neutrophil axis (i.e. PSGL-1) is a promising and effective approach to prevent thrombosis in the context of inflammation and infection, highlighting the significance of the interaction in the development of thrombosis during sepsis.