Targeting Neutrophil Extracellular Trap under Flow in Patients with Immune-Mediated Thrombotic Thrombocytopenic Purpura.

Neutrophil NETosis is a unique form of cell death, characterized by release of decondensed chromatin and anti-microbial contents to the extracellular space, involved in infection, inflammation, and thrombosis. However, the role of NETosis in pathogenesis of immune-mediated thrombotic thrombocytopenic purpura (iTTP) and how a targeted therapy affects the accumulation of neutrophil extracellular traps (NETs) under flow remain to be determined. Flow cytometry demonstrated that the percentage of neutrophils undergoing NETosis in whole blood from iTTP patients on admission is significantly increased with a concurrent decrease in the capacity of inducible NETosis by shigatoxin. Following therapy, the percentage of H3Cit+MPO+ neutrophils was significantly reduced with an improvement of inducible NETosis in these patients. Additionally, little to no NET and thrombus formation was detected under flow in whole blood from iTTP patients when platelet counts were very low, but the NET and thrombus formation were dramatically increased following therapy when platelet count rose to 50x109/L or was restored to normal with donor platelets. Similarly, there was no thrombus and NET accumulation under flow in whole blood from vwf-/- mice, but NET accumulation was significantly higher in Adamts13-/- than wild-type mice. Finally, recombinant ADAMTS13 or caplacizumab (or anfibatide) prevents the NET and thrombus formation under flow in whole blood from iTTP patients (or Adamts13-/- mice). These results indicate that neutrophil NETosis and NET formation depend on platelets and VWF in iTTP, and a targeted therapy such as recombinant ADAMTS13 or caplacizumab may prevent the NET and thrombus formation under flow in iTTP.