OC 46.2 - Fibrin Film Formation is Enhanced by Red Blood Cells and Reduces Platelet Spreading

Background: Thrombi are heterogeneous, consisting of fibrin, red blood cells (RBCs), and platelets. Recent studies show that fibrin films develop on the surface of clots, playing a protective role. Evidence is emerging that fibrin films are also present in intravascular thrombi; however, little is known about the role of these films and their interactions with blood cells and the vasculature.

Aims: Investigate the effect of haematocrit on fibrin films, the role of fibrin films in platelet spreading and the impact of inflamed endothelial cells on fibrin film formation.

Methods: Blood was obtained from healthy volunteers. Haematocrit was adjusted to 10, 30, 50, 70% by dilution with autologous plasma. Effects on clot film coverage was investigated by scanning electron microscopy. Platelets spreading on clot surfaces in the presence or absence of film was investigated by laser scanning confocal microscopy (LSCM). Human umbilical vein endothelial cells (HUVECs) were stimulated with lipopolysaccharide (LPS), interleukin-1β (IL-1β) or tumour necrosis factor-α (TNFα). Inflammation markers, including monocyte chemoattractant protein-1 (MCP-1) and tissue factor, were analysed by enzyme-linked immunoassay. Effects of HUVEC inflammation on fibrin film was analysed by LSCM.

Results: Blood clots throughout the haematocrit range developed fibrin film; however, clots with 10% haematocrit showed less film coverage than those with a higher haematocrit. Spreading of platelets on clots with film was reduced compared to clots without film or fibrin monomer layer. MCP-1 level was high in all treated HUVECS whilst tissue factor level was high in TNFα treated cells. There were no significant differences in film thickness in clots made with purified fibrinogen on the HUVECS with or without inflammation.

Conclusion(s): Our data show that RBCs support fibrin film coverage on clots, and that fibrin films impede platelet spreading. These findings may have important implications for boundary formation in clots and thrombi.