In vivo fibrin clot retraction occurs in the needle-injury model. (A) Images depict a 3-dimensional (3D) surface rendering of a representative thrombus, forming as a result of the needle-injury model in vivo. The surface of the platelet thrombus is in red (Dylight 649 anti-mouse GP1ba antibody), the surface of the fibrin network is in white (Alexa Fluor 546 anti-fibrin antibody), and the surface of collagen in the basement membrane is in green (autofluorescence). Supplemental Video 1 provides an animated perspective of this same thrombus and orthogonally cuts through the thrombus to highlight the underlying ring-shaped fibrin network, which is the key feature of the needle-injury model. (B) The thrombus size (red; left y-axis) and the area occupied by the fibrin network (green; left y-axis) were quantified (as described in “Methods”) and expressed as a percentage of thrombus size at 5 minutes. The total fibrin amount (blue; right y-axis) was also quantified (as described in “Methods”) and expressed as a percentage of the total fibrin amount at 5 minutes. The graph depicts the mean ± standard error of the mean (SEM). (n = 8); ***P < .001; ****P < .0001 by 2-way analysis of variance (ANOVA) relative to value at 5 minutes. (C) Confocal maximum-intensity projection of the ring-shaped fibrin network (detected via Alexa Fluor 546 anti-fibrin antibody), which reduces in size over the first 2 hours of thrombus lifetime. The superimposed yellow line outlines the fibrin network perimeter at 5 minutes. (D) A confocal maximum-intensity projection of a representative thrombus with platelets (Dylight 649 anti-mouse GP1bα antibody) and the plasminogenFITC-bound fibrin network both reducing in size over the first 2 hours of thrombus lifetime. The superimposed yellow lines show the periphery of the thrombus and its underlying fibrin network at 5 minutes. (A-B,D) The direction of blood flow (arrow) and scale bars (10 μm) are shown.
