CLSM analysis of thrombi generated on a collagen surface in a healthy control and type 2B patients at a 1500 s−1shear rate.
Experimental conditions were as described in the legend to Figure 2except that only a typical high shear rate (1500 s−1) was applied. Thrombi generated at 7 minutes of perfusion of blood from a healthy control and selected type 2B VWD patients (2B-3, -4) were fixed before viewing by CLSM. Surface coverage (top panel), maximum height (middle panel), and total volume (bottom panel) of thrombi within a defined area (211 × 317 μm each) were evaluated. Data represent the mean + SD of 5 areas randomly selected in each single perfusion. One-way factorial ANOVA and the Scheffé method were used for analysis of variance and for comparisons with controls, respectively, with assistance of Stat View computer software (Abacus Concepts, Berkeley, CA). Asterisks indicate statistically significant differences from respective controls (P < .01). Statistical analyses demonstrated that the height and volume of type 2B VWD thrombi were about half those of a healthy control, whereas surface coverage was comparable to normal (left panels). The reduced thrombus height and volume in type 2B VWD were completely normalized when purified VWF (20 μg/mL) was added to type 2B VWD blood prior to perfusion. The height and volume of type 2B VWD thrombi were significantly reduced compared with a healthy control, even when the collagen-coated surface was incubated with 200 μL purified normal VWF (20 μg/mL) for 1 hour prior to perfusion (right panels).
