Abstract 14208: Annexin A5 Inhibits Endothelial Inflammation Induced by Lipopolysaccharide-Activated Platelets and Extracellular Vesicles

Abstract

Introduction: Sepsis is a leading cause of death worldwide with no targeted therapeutics available. Sepsis results from a dysregulated immune and inflammatory response. During sepsis or lipopolysaccharide (LPS) stimulation, platelets are activated, and release procoagulant/proinflammatory membrane-derived extracellular vehicles (EVs). Our previous studies show recombinant human annexin A5 (Anx5), a phosphatidylserine-binding protein, inhibits inflammation and improves survival in endotoxic mice, a model of sepsis. Hypothesis: Anx5 will block the pro-inflammatory response induced by LPS-activated platelets and EVs in vascular endothelial cells (ECs). Methods: ECs were cultured from adult C57BL/6 mice. Platelets and EVs were activated by LPS and added to ECs for four-hours with/without AnxA5. Inflammatory cytokines and adhesion molecule expression was quantified in ECs using RT-qPCR. Endothelial monolayer structural integrity was measured in septic conditions using trans-endothelial electrical resistance (TEER). Monocytes were added to LPS-treated ECs and adhesion was quantified via brightfield microscopy. Results: Supplementation of annexin A5 (1 μg/ml) on ECs treated with LPS-activated platelets and EVs reduced the expression of inflammatory cytokines and adhesion molecules on ECs when compared to ECs treated with LPS (1 μg/ml) activated platelets and EVs alone. In septic conditions, endothelial monolayers that were treated with annexin A5 had a higher TEER compared to the monolayers without annexin A5, indicating that annexin A5 preserved endothelial monolayer structural integrity in septic conditions. Annexin A5 treated ECs also had significantly less monocytes adhesion in septic conditions when compared to ECs treated without annexin A5. Conclusions: The proinflammatory response induced by activated platelets and EVs is inhibited by Anx5. The ability to maintain the structural integrity of endothelial monolayers and reduce monocyte adhesion during septic conditions, makes Anx5 a promising treatment option for sepsis.