Impaired hemostasis and protection against thrombosis in protease-activated receptor 4-deficient mice is due to lack of thrombin signaling in platelets

Platelets from protease-activated receptor 4 (PAR4)-deficient mice are unresponsive to thrombin, and Par4-/- mice have prolonged bleeding times and are protected against thrombosis. However, in addition to its role in platelets, PAR4 contributes to thrombin signaling in cells in the blood vessel wall that might participate in hemostasis and thrombosis, such as endothelial cells. To determine whether the hemostatic and thrombotic phenotypes of Par4-/- mice were due to loss of PAR4 function in hematopoietic vs. other cell types, tail bleed times and thromboplastin-induced pulmonary embolism were examined in lethally irradiated mice reconstituted with Par4+/+ or Par4-/- bone marrow. In Par4+/+ and Par4-/- mice reconstituted with Par4+/+ marrow, the median tail bleed times were 2.0 and 1.7 min, respectively, vs. >10 min for both Par4+/+ and Par4-/- mice reconstituted with Par4-/- marrow. In the pulmonary embolism model, Par4+/+ and N4-/- mice reconstituted with Par4+/+ marrow survived a median of 3.7 and 2.8 min, respectively, after administration of thromboplastin, vs. >20 min for both Par4+/+ and Par4-/- mice reconstituted with Par4-/- marrow. Further, the phenotype of mice reconstituted with N4-/- marrow was almost as dramatic as that seen in Nf-e2-/- mice, which lack platelets. These data strongly suggest that increased tail bleed times and protection against thrombosis in Par4-/- mice are accounted for by lack of PAR4 function in platelets, emphasize the importance of thrombin signaling in platelets among the multiple pathways and cell types that govern hemostasis and thrombosis.