Platelets mediate fibrin-induced thromboinflammation and NETosis through a GPVI-polyphosphate-FXIIa procoagulant axis.

Fibrin deposits drive thromboinflammation and neutrophil extracellular trap (NET) formation (NETosis) in various pathophysiological situations, but the underlying mechanisms remain unclear. Similarly, it is unknown how, irrespective of diabetes, hyperglycemia exacerbates thromboinflammation. Using a microfluidic model of thromboinflammation based on the perfusion of partially recalcified citrated human blood at a venous flow rate over immobilized fibrin, we investigated these mechanisms. Blood was stained to detect P-selectin, phosphatidylserine, polyphosphates, fibrin, neutrophils, and NETs. The roles of glycoprotein VI (GPVI), integrin α2bβ3, polyphosphates, factor XIIa (FXIIa), tissue factor, thrombin, fibrin polymerization, glucose, and glucose uptake were explored using specific inhibitors and by spiking blood with glucose. Perfusion of partially recalcified blood over a fibrin monolayer consistently triggered a platelet-dependent thromboinflammatory cascade. Pharmacological inhibition and analysis of activation at the individual platelet level revealed that this cascade was initiated by a subpopulation of procoagulant platelets, which activated second-responder platelets via GPVI-, FXIIa-, and polyphosphate-dependent thrombin generation, leading to fibrin polymerization, and neutrophil recruitment. This sequence was maintained but exacerbated in hyperglycemia, drastically increasing intrathrombus neutrophil recruitment and NETosis, the latter prevented by inhibition of glucose transporters. Our results highlight the importance of a GPVI-FXIIa-polyphosphate axis in mediating thromboinflammation and NETosis when flowing human blood encounters immobilized fibrin and the interest of targeting this axis and glucose transporters to prevent thromboinflammation and its exacerbation by hyperglycemia.