Histones link inflammation and thrombosis through the induction of Weibel–Palade body exocytosis

Abstract

Essentials• Dysregulated DNA and histone release can promote pathological immunothrombosis.• Weibel-Palade bodies (WPBs) are sentinel-like organelles that respond to proinflammatory stimuli.• Histones induce WPB exocytosis in a caspase, calcium and charge-dependent mechanism.• A targetable axis may exist between DNA/histones and WPBs in inflammation and immunothrombosis.Summary. Background: Damage-associated molecular patterns (DAMPs), including molecules such as DNA and histones, are released into the blood following cell death. DAMPs promote a procoagulant phenotype through enhancement of thrombin generation and platelet activation, thereby contributing to immunothrombosis. Weibel-Palade bodies (WPBs) are dynamic endothelial cell organelles that contain procoagulant and proinflammatory mediators, such as von Willebrand factor (VWF), and are released in response to cell stresses. VWF mediates platelet adhesion and aggregation, and has been implicated as a procoagulant component of the innate immune response. Objective: To determine the influence of histones and DNA on WPB release, and characterize their association in models of inflammation. Methods:We treated C57BL/6J mice and cultured endothelial cells with histones (unfractionated, lysine-rich or arginine-rich) and DNA, and measured WPB exocytosis. We used inhibitors to determine a mechanism of histone-induced WPB release in vitro. We characterized the release of DAMPs and WPBs in response to acute and chronic inflammation in human and murine models. Results and conclusions:Histones, but not DNA, induced the release of VWF (1.46-fold) from WBPs and caused thrombocytopenia (0.74-fold), which impaired arterial thrombus formation in mice. Histones induced WPB release from endothelial cells in a caspase-dependent, calcium-dependent and charge-dependent manner, and promoted platelet capture in a flow chamber model of VWF-platelet string formation. The levels of DAMPs and WPB-released proteins were elevated during inflammation, and were positively correlated in chronic inflammation. These studies showed that DAMPs can regulate the function and level of VWF by inducing its release from endothelial WPBs. This DAMP-WPB axis may propagate immunothrombosis associated with inflammation.

Platelet proteome reveals novel pathways of platelet activation and platelet-mediated immunoregulation in dengue

“…It has been previously shown that free histones bind to and activate platelets in vitro and in vivo [24,25,26,27]. However, the role played by circulating free histones in platelet activation during dengue infection remains unclear.…”

“…Treatment of platelets from healthy uninfected donors with recombinant human histone H2A significantly increased P-selectin translocation to the surface ( Fig 6A ) and secretion of PF4/CXCL4 into the supernatant ( Fig 6B ). Unfractionated histones have been shown to activate cellular responses through mechanisms involving toll like receptor (TLR) binding and calcium-mediated signaling [24,25,26,28,29]. Treatment of platelets with the calcium chelator BAPTA-AM (20 μM) significantly impaired P-selectin surface expression and PF4 secretion by histone H2A-activated platelets ( Fig 6C and 6D ).…”

“…Similarly, when whole blood is exposed to histones in vitro , histones bind to platelets leading to platelet aggregation [26]. Accordingly, injection of histones into mice leads to histone accumulation in sites of thrombosis and to thrombocytopenia [24,26,27]. In experimental sepsis in mice and in patients with sepsis, a syndrome with many parallels with severe dengue, circulating histones are major mediators of vascular damage and disseminated intravascular coagulation (DIC) [27,29,43].…”

“…In experimental sepsis in mice and in patients with sepsis, a syndrome with many parallels with severe dengue, circulating histones are major mediators of vascular damage and disseminated intravascular coagulation (DIC) [27,29,43]. In addition to platelet activation, circulating free histones also activate endothelial cells amplifying the activation of inflammation and coagulation through endothelium expression of tissue factor and extrusion of Weibel-Paled bodies [24,28]. While the roles played by cell-free histones in vasculopathy, shock and organ dysfunction in dengue remain to be precisely determined, our observations strongly suggest that histone-mediated platelet activation may contribute to dengue pathogenesis.…”

Parasite histones mediate leak and coagulopathy in cerebral malaria

“…In vitro , histone binding to the EC membrane causes toxicity and barrier disruption (22, 23). The cationic domain of histones also induces Weibel Palade body exocytosis, endothelial activation and thrombocytopenia through platelet aggregation on von Willebrand Factor strings (25). Histones further induce a procoagulant phenotype through upregulation of endothelial tissue factor (26).…”