Figure 2. Perivalvular venous endothelium is resistant to leukocyte rolling and thrombus formation.

(A) Spontaneous leukocyte rolling at the saphenous venous valve was observed and quantitated in PROX1-GFP mice using rhodamine 6G. Arrow indicates the direction of flow, and arrowheads indicate individual leukocytes adherent to the vessel wall. Dashed lines outline the valve sinus region. The number of rolling leukocytes per millimeter vessel length was measured upstream of the valve (US), in the valve and valve sinus regions (V/S), and downstream of the valve (DS) (right) (n = 5 valves imaged in 3 mice). (B) Thrombus formation was stimulated in PROX1-GFP animals around the saphenous venous valve by application of extravascular thrombin, and clot visualized by accumulation of rhodamine 6G–positive platelets and leukocytes (orange) at 120 seconds (left). The percentage of vessel area covered by thrombus was measured upstream of the valve (US), in the valve and valve sinus regions (V/S), and downstream of the valve (DS) (right) (n = 8 valves imaged in 4 mice). (C) Visualization of thrombus formation over the saphenous venous valve of a PROX1-GFP mouse using anti-fibrin antibodies 10 minutes after thrombin application. The images are representative of those obtained from analysis of 3 separate experiments. White dashed lines indicate luminal venous endothelial cells, and green dashed lines indicate perivalvular endothelial cells. For each graph the mean is shown as the value of the bar with dots representing each replicate, and error bars indicate SD. Significance was determined by a paired 2-tailed t test and corrected for multiple comparisons. *P < 0.025; **P < 0.01.