Figure 3. Thrombin triggers protracted and lethal pulmonary thrombosis in mice.

WT mice were administered IV with 250 U/kg (n = 5 mice) or 500 U/kg (n = 4 mice) thrombin, and pulmonary circulation was imaged using qFILM. (A and B) qFILM images of the same FOV at different time points are shown. t = 0 seconds (s) corresponds to time point before and t > 0 s correspond to time points following IV thrombin administration. Pulmonary thrombosis was absent at t = 0 s. Platelets are shown in green and pulmonary microcirculation in purple. (A) Following 250 U/kg IV thrombin, small (<500 μm²) and medium (500–1000 μm²) platelet-rich thrombi (white arrowheads) sequestered in the pulmonary arteriole (t = 10–12 s) and obstructed blood flow (t = 20 s). (B) Following 500 U/kg IV thrombin, small (<500 μm²) and medium (500–1000 μm²) platelet-rich thrombi (white arrowheads) sequestered in the pulmonary arteriole to occlude the arteriolar bottlenecks. The mouse died by t = 3 minutes, leading to arrest of pulmonary blood flow, which was evident by the reduced intensity of vascular dye (purple fluorescence) and stationary erythrocytes (Supplemental Video 5). Asterisks denote alveoli. White arrow mark the direction of blood flow. The diameters of the arterioles shown in A and B are 39 μm and 44 μm, respectively. Complete qFILM time series corresponding to A and B are shown in Supplemental Videos 4 and 5, respectively. (C and D) Pulmonary thrombi area plotted as a function of time following 250 U/kg (C) and 500 U/kg (D) IV thrombin within FOVs shown in A and B, respectively. Red and black arrows indicate pulmonary thrombi maximum area values and the time of mouse death following 500 U/kg IV thrombin, respectively. (E) Survival rate during qFILM experiments in WT mice IV administered with either 250 U/kg (n = 5 mice) or 500 U/kg (n = 4) thrombin (P = 0.046, log-rank test). (F) Pulmonary thrombi max areas in mice following 250 U/kg (n = 5 mice) and 500 U/kg (n = 4 mice) IV thrombin. Pulmonary thrombi max areas were compared using Wilcoxon-Mann-Whitney test. Data are shown as mean ± SEM. (G) Three-dimensional qFILM image of a lethal pulmonary thrombosis developed within a large pulmonary arteriole (57 μm) of a mouse administered with 500 U/kg IV thrombin. Platelets (green) and pulmonary microcirculation (purple). Refer to Supplemental Video 6. Scale bar: 50 μm.