The Syk inhibitor BI 1002494 impairs thrombus infill in a murine femoral artery occlusion without affecting hemostasis.

Knowing structural organization of a clot at the single-cell level could lead to the development of drugs targeting specific structural features. We tested this premise in a murine femoral artery occlusion model using a narrow ferric chloride (FeCl3) application strip to limit induction intensity. Under these conditions, occlusive clot formation was sensitive to the spleen tyrosine kinase (Syk) inhibitor BI 1002494, an indication of normative platelet response. Samples perpendicular or longitudinal to blood flow were imaged by montaged electron microscopy. Platelets were the predominant cell type. Tightly packed platelets were anchored to FeCl3-induced damaged portions of the vessel wall, and aggregates of tightly packed platelets extended inward. Overall, the clots had a structure in which loosely packed platelets, often discoid in shape and rich in α-granules, filled pockets within the clot surrounded by zones of tightly adherent platelets. Red blood cells (RBCs), mainly entrapped, squeezed, and polyhedral in shape, were distributed in scattered patches. Based on platelet morphology, any effect of RBCs on platelet activation extended for a short distance, ∼5 μm. In Syk inhibitor-treated mice, structural formation of an occlusion was strongly inhibited; infill was impaired, resulting in a highly porous clot rich in dispersedly aggregated discoid shaped platelets. Blood flow was normal, and inhibitor had no apparent effect on the structure of a femoral puncture wound clot. We suggest that BI 1002494 produced a selective inhibition of thrombus structure by depressing intraplatelet signaling below a crucial threshold in the high flow occlusion model, but not in the lower flow puncture wound model.